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Cardio2D Manual - Multi Channel Systems

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1. 7 USB MEA2 56 2 22 7 256 9wellMEA 56 256 aue 56 5 5 5 7 56 Start Stop 52 Stimulation 36 Inter Pulse Interval 36 P lse Shape iil tiae 36 deu 36 SV OU 36 Voltage Pulse dv bec Badia ons 36 Stimulation with MEA2100 System 39 Clear Stimuulus sus 39 Complete Stimulus Pattern 39 Define Stimulus tro ies 39 Select Stimulus 39 Single Stimulus pattern 39 Stimulus Pattern 39 BIOIOGICA M 39 Flat LIWeu 39 Marker Signal iiid 39 ee P 39 39 SING MOVE 39 Stimulus Generator 36 Summary Window 53 Multi Well Multi MEAS 53 T Technical Support 121 Test Model Probe 121 Theoretical Background 119 Timestamp 63 Troubleshooting 117 V Velocity 101 Viability 29 125
2. The HB Averager tab is divided in three separate windows The MEA electrode field the Recording overlay and the Channel overlay window 105 Cardio2D Manual Important Please pay attention to the Analyze dialog To use the heartbeat average tool you have to enable the check box in the HB Averager tab of the Analyze dialog in main menu Tools Analyze X HB Detection HBAwerage zp On Recording Fre HB Interval 50 ms Post HB Interval 500 m rs Apply to all Recordings MEA Electrode Field Recorded data traces are displayed in the MEA electrode field To adapt the y axis please use the drop down menu The MEA electrode field shows the averaged heartbeats of all channels of the currently selected recording The MEA electrode field corresponds with the left overlay plot window Recording Overlay The data traces of the MEA electrodes selected in the MEA electrode field and highlighted in white are displayed in the overlay plot window in different colors Recording Overlay Hec 00 41 Rec 01 41 Rec 02 18 5 o LL 50 Time ms Tee f 9 8 In the Recording Overlay window the heartbeats of the different recording phases of one single channel are displayed in an overlay plot See the number of the channel in the upper left edge of the plot in this example channel number 33 To select this channel please select the
3. 1 2 3 B 4 10 Recording Sequence 05 1 B 3 4 5 n 9 10 11 0 Analyze 04 100 Time s 100 uw This tabbed page lists the single recording sequences from one electrode Select the electrode in the Channel control field of the Control Window Via ROI you can discard one part or several parts of the recording sequence which are now excluded from the analysis Additionally it is possible to discard a complete recording sequence by deselecting the respective check box The data in the ROI will not be included in the analysis Select Heartbeat tab Please see the picture below The single channel view on top of the tab page shows amplitude and rate of the heartbeats of one recording phase The MEA electrode window corresponds directly with the color map The field potential of that single electrode is shown in MEA electrode window which is selected via mouse in the color map Select a plot of Latency or a plot of Velocity from the drop down menu in the Latency window 73 Cardio2D Manual The color map corresponds with the single channel view and shows the excitation patterns of that heartbeat marked with the blue arrow in single channel view Field Potential Selection Heartbeat Average HT Analysis Single Channel View MNA S REESE NCC IE RE 33124 33126 33128 33 13 EE 3313 MEA Electrode 10 d R
4. 10 0 500 0 200 400 Time ms Time ms Test only 100p i 100p d Select QT related Analysis tab Please see the picture below Important Please pay attention to the Analyze dialog To use the QT related Analysis tool you have to enable the check box in the HB Average tab of the Analyze dialog in main menu Tools and the respective check box in the QT related Analysis tab analyze x HE Detection HB amp verager 20 HT wl On Detection Deadtime mz Slope Calculation 10 0 ms 75 Cardio2D Manual Select one channel from the Heartbeat window pane of the Control Window and the desired channel will be displayed with its different recording phases in the QT related Analysis window on the left side If possible a dose responds curve of this channel will be calculated and displayed in the QT like Interval window of the right side You can choose whether you want to display a normalized or an absolute curve from the drop down menu Field Potential ROI Selection Heartbeat OT Analysis HT Interval ms Dose Response 100 1000 10000 100000 Dose ph Absolute QT related Interval 400 76 Theoretical Background Main Menu Fil Tools Help File ES CardiozD Open Raw Data Open Result File Save Result File Exit Menu to open or sa
5. The pulse shape of a stimulus pulse is usually biphasic symmetric rectangular The parameters are the negative or positive sign of the first pulse phase the duration of each phase and the amplitude of the pulse Click the check box Symmetric and the pulse will automatically be symmetric and biphasic To design the pulse shapes please use the numeric up down controls Phase 1 and Phase 2 The IPI the inter pulse interval is the time frame between two pulses in sequence The red line indicates the stimulus pulse the blue line indicates the TTL blanking pulse Stimulation Pulse Channel 1 Inter Pulse Interval TTL Pulse for Blanking Signal from Sync Out Sync Out Channel 1 Extension Extension Blanking Pulse The Sync Out menu is for the blanking feature if an amplifier with blanking circuit is connected Synce On m E tensian 300 36 Cardio 2D Software When using the blanking feature please enable the Sync Out check box You have to define the time of Extension The TTL pulse for the blanking signal starts synchronal to the stimulus pulse but it should last longer to avoid stimulus artefacts This time interval is called extension time The extension time varies between 0 and 1000 us the default is 300 us In some cases it may be necessary to see the stimulus artefacts Please deselect the check box Sync Out to switch off the Sync Out signal Additional plea
6. 27 Control Window Cardio2D 27 Control Window Cardio2D 89 D Detection Parameters 71 E Error messages 117 F Field Potential Cardio2D 92 Channel overlay 92 Color MaDe peia 92 MEA electrode field 92 Single channel view 10 Field Potential Duration 112 H Hardware Acquisition System 10 MC Card 10 MEA 2100 System 10 USB ME32 FAl System 10 USB MEA256 System 10 USB ME Systems eese 10 HB Averager 109 Channel overlay 109 MEA electrode field 109 Recording overlay 109 Heartbeat Cardio2D 101 Color mapas a PO 101 Latency Velocity 101 MEA electrode 101 Single channel 101 Heartbeat window 17 L Labbook 30 Latency window 17 61 Te Dao podria RED d 61 License Key 7 Log file window 17 53 Main menu 17 Experiment 17 TT 17 17 MEAS iso eal abe 17 SELOS neria 17 MEA Electrodes 17 56 Analyzer 95 56 VJ dep ER EE EE 56 Propagation 56 AmpHt de 5 qnt ie se 56
7. 75 QRS ST Interval Interval Interval QT Interval Theoretical Background Reference Estimation of Action Potential Changes from Field Potential Recordings in Multicellular Mouse Cardiac Myocyte Cultures Marcel D Halbach et al Cell Physiol Biochem 2003 13 271284 117 9 1 9 2 Appendix Technical Support Please read the Troubleshooting part of the user manual or Help first Most problems are caused by minor handling errors Contact your local retailer immediately if the cause of the trouble remains unclear Please understand that information on your hardware and software configuration is necessary to analyze and finally solve the problem you encounter If you have any question or if any problem occurs that is not mentioned in this documentation please contact your local retailer The highly qualified stuff will be glad to help you Please keep information on the following at hand Description of the error the error message text or any other useful information and of the context in which the error occurred The more information on the actual situation you can provide the easier it is to track the problem The serial number of the device You will find it on the bottom side of the main unit and on the amplifiers The software and hardware version you are currently using On the Help menu click About The displayed dialog box shows the version numbers The operating system and service p
8. EL ES b bl EH 5 jot pot jo joe pe fot pol 1 edt BEHENBGEHESEHESHEHESSEEHHHEEERNN ELE Lll ted ded dol def bod Jo 15 ui dz FE TET ITTE TETT ALLL m LE mei 720 Ae te Hi ELT ELLE Ed E aa A er ot AA fzr at er El ol ot tet ip tol fet te flay fal 141 351 143 er a Ga 24 Cardio 2D Software Click View Summary Page to open the Summary window This window is only available if more than one MEA at a time is connected The window presents an overview and all connected MEAs are visible The dark green color code spotlights the MEA which is displayed in the MEA tabbed page The layout of the MEA is displayed respectively ES cardio2D File Experiment Tools Settings Help Summary MEAZ 1 MEA 2 MEA 3 MEA 4 lt 07 32 55 Cardio2D start lt 07 32 58 gt Initialize MEA 1 07 32 58 Initialize MEA 2 lt 07 32 59 gt Initialize MEA 3 lt 07 32 59 gt Initializ
9. The settings in this tool bar influence the complete sequence of the stimulus patterns Amplitude 95 0 55 Offset mV 0 oo x Digital Out off Modulate the Amplitude and the Offset mV of the complete pattern via up down boxes Clicking the Loop 99 button enables you to infinitely repeat the defined stimulus pattern Load a previous created stimulus pattern save or delete the pattern When using marker signals you have to define which one of the Digital Out ports should show the corresponding TTL signals Select one of the marker ports from the drop down menu and connect a TTL signal source to the port Download and start the created stimulus pattern via the Download icon If downloading Is not necessary the icon is not available This way you have an optical feedback for the download 48 4 9 Cardio 2D Software Analyzer Settings Click Analyze icon The Analyzer Settings dialog will open Click HB Detection tab As in the Labbook settings the same is in Analyzer settings You can have local settings on each MEA or each well of a MEA or you control all MEAs at a time via global settings The global settings white background overwrite the local settings orange background analyzer Settings x HE Detection Averager 20 Minimum Time 200 us I asimum Rise Time FOO ps C Fixed Amplitude Signal Dependent
10. 129046 0 794 1259700 In the header of the comma separated value file all exported parameters listed The square brackets show the respective units Import the ASCII result file into software of your choice for further analysis 111 Cardio2D Manual Heartbeat tab Result Export Peak Select Recording Recording 0 no dose specified Maximum Minimum Peak4o Peak Amplitude Inter Beat Interval Ed 2 a amp a 82 Peak to Peak Duration 23 43 59 7 Slope Ge qe No Sere 288666080 c 16 26 36 46 86 io 5 6 87 Export Heart Beats The Heartbeat tab provides parameters which characterize the depolarization and beat rate Heartbeat 2D tab Result Export E xj Heartbeat Heartbeat 2D Averaged Heartbeat Dose Response Dose Response 2D Select Recording Recording 0 dose spected 20 97 Data Latency Velocity Number of Active Electrodes First Electrode Last Electrode Blectrode Latencies m 2 98 amp 92 Export 2D Heart Beat Data The Heartbeat 2D tab provides parameters which characterize 2D signal propagation across the MEA Averaged Heartbeat tab Result Export E xl Averaged Heartbeat Dose Response
11. D ater AUS MES c r eet U ph s 100 nh s FENEINET s T 100 s In the recording control window the name s of the currently analyzed Cardio2D raw data and or result file is displayed Additional the information about the recordings saved in this experiment are listed in a table To select one recording phase to be analyzed please click onto the desired row which will be highlighted in dark blue and displayed in the single channel view of the MEA window To view raw data and results of a recording phase in the MEA window please click onto the desired row in the table The currently selected recording phase will be highlighted in dark blue The table has five columns the ID of the recording phase the Type of the phase was it an apply A or a test phase T The Dose of the recording phase with its unit the Compound and the Duration Channel control of Cardio2D The channel control window is similar build to the heartbeat window in Cardio2D but the function is different It also shows the MEA electrodes in the electrode grid You can zoom to any electrode by clicking on the icon Show Channel Data and the electrode The zoomed electrode is displayed in the larger single channel view 85 6 3 Cardio2D Manual In this screenshot a MEA electrode grid 8 x 8 is displayed Different layout grids are possible depending on the selec
12. Latency 103 Cardio2D Manual Select Velocity from the drop down menu Recording 0 Recording 1 Recording 2 Recording 3 Recording 4 Recording 5 Recording 9 E E E S EA D E T mm Time s Velocity E The diagram shows the velocity cm s of the excitation in dependency of the time s between the minimum and the maximum color coded in overlay for each recording separately This Latency and Velocity windows are directly linked to the Single channel view the Color Map and the file information table in the Control Window Click on one of the data points and the mouse pointer turns into a hand The blue arrow in the Single Channel View will mark the corresponding heartbeat and the color map will change immediately and show the excitation pattern at that actual time If you click any data point from a different recording different color additionally the respective recording will be highlighted in blue in the file information table 104 Theoretical Background 6 7 HB Averager Click the HB Averager tab Field Potential Selection Heartbeat HB amp verager HT Analysis 01 41 21 33 Rec 05 41 31 33 05 20 41 33 62 33 51 33 Recording Overlay Channel Overlay 10 ADD A 0 500 0 200 400 Time ms Time ms Testonly 100p i 100p E
13. Settings Data Acquisition File Locations View Summary Page Menu to define the data acquisition device and to determine the path for the data and log files 22 Cardio 2D Software Click Data Acquisition in the Settings menu The Data Acquisition Settings dialog opens This dialog is the same which shows up before starting Cardio2D for the first time Please see above ES Data Acquisition Settings X Device USB ME 255 v Sample Rate 4096 mv Driver Version 1 MEA 8 8 100 10 MEA 2 MEA 8 8 100 10 MEA 3 MEA 8 8 100 10 MEA 4 MEA 8 8 100 10 Close Please enter the drop down menus to select the current data acquisition In dependency of the data acquisition device you can modify the Sample Rate and the Range The Resolution Number of Channel and the Driver Version will be displayed Define the type of MEAs used in the experiment Click File Location in the Settings menu The Location Settings dialog opens ga Location Settings x Haw Data CA Dokumente und Einstellungen sCardia2D D ata Results CA Dokumente und Einstellungen ardia2D D ata Lag Files CA Dokumente und Einstellungen ardiazD ata ER Close Please define the file path for the raw data and the corresponding setting files the result and the log files When using an USB ME256 device for data acquisition it is possible to connect up to fou
14. 10 StdDey Detection Deadtime 200 ms Min Max Global Min Mas Search 10 HB Timestamp b inimum In this dialog you define the parameter for detecting the waveform of a field potential Please see also Automated Waveform Detection and Detection Parameter The feature Comparing Local and Global Settings is also available for this dialog Please read chapter Labbook The first parameter for detection is the Rise Time Select the minimum and the maximum of the rise time from the drop down menus The range involves 100 us to 2 ms for the minimum rise time and 200 us to 5 ms for the maximum rise time Next parameter is the Amplitude You can decide to have a fixed amplitude for detection or a signal dependent amplitude The fixed amplitude involves a voltage range from 10 uV up to 1 mV Using the signal dependent amplitude the waveform detection is more flexible The detection routine detects the amplitude in relationship to the background noise The amplitude is calculated by the standard deviation of the signal times the factor you can choose It is recommended to use a signal dependent amplitude The following parameters characterize the detected field potential Detection Deadtime defines the time in which no detection is performed 49 Cardio2D Manual The drop down menu Min Max Type describes the type of the minimum or maximum Local or Global These options refer to one single field
15. 5 1 5 2 5 3 Table of Contents Introduction About this Manual Welcome to Cardio2D and Cardio2D Important Information and Instructions Important Safety Advice Guarantee and Liability Operator s Obligations Terms of Use for the Program Limitation of Liability Software and Hardware Installation Installing the Cardio2D Software First Use of Cardio2D after Installation Data Acquisition Hardware Cardio 2D Software The Cardio2D Program Overview Control Window Control View Viability Labbook Compound Basic Stimulation of MEA Electrodes Stimulation with MEA2100 System Analyzer Settings Start Stop Log File Summary Window MEA Window MEA Electrodes Single Channel View Latency Velocity Window Color Map General Software Features Experimental Proceeding General Performance Noise Level Detection Parameter Automated Waveform Detection _ 9 Uu N 19 19 27 28 29 29 31 35 39 49 52 52 53 54 55 56 58 60 64 67 67 68 69 Cardio2D Manual 6 1 6 2 6 3 6 4 6 5 6 6 6 7 6 8 6 9 7 1 7 2 Cardio 2D Analyzer Software The Cardio2D Analyzer Program Overview Control Window Cardio2D MEA Window Cardio2D Field Potential Cardio2D Selection Heartbeat Cardio2D HB Averager Analysis of Field Potential Duration Export of Results Troubleshooting Troubleshooting Error Messages Theoretical Background Cardiac Action Potential A
16. Dose Response 2D Averaged Heart Beat Intervals Select Recording Recording 0 no dose specified veraged Heart Beat Intervals Average Std Dev e eee de Ge de e G5 3 5 m 03 4 m eom SESDE m a 80 oe Export Averaged Heartbeat Interval The Averaged Heartbeat tab allows you to export averaged heartbeat intervals 112 Dose Response tab B Result Export Heartbeat Heartbeat 2D Averaged Heartbeat Dose Response Dose Response 2D NaPeak from Raw Data Peak to Peak Amplitude PeaktoPeak Duration PeaktoPeak Slope Max Slope m Select Recording Recording 0 ino dose specified o Heartbeats Inter Beat Interval Beat Rate QT from Averages Field Potential Duration 2 98 8 T 8 8S 2 28 8 28 82 Repolarization Amplitude Export Dose Response Results Theoretical Background The Dose Response tab provides parameters which indicate the effect of different doses of a compound on heartbeat characteristics with focus on a single electrode Dose Response 2D tab B Result Export E xl Heartbeat Heartbeat 2D Averaged Heartbeat Dose Response Dose Response 2D 2D Signal Propagation Latency Velocity Number of Active Electrodes Yv Select Recording Recording 0 dose specified Export 2D Dose Response Results The Dose Response 2D t
17. If you select Heart Rate Hz the rate of the heartbeats is calculated from the reciprocal value of the RR interval The last actual value of the RRI ms or of the heart rate Hz is displayed in red above the plot RRI ms 626 E oc Lc u i t etta tt IE REI NE RSS NEUE 10 15 Zh Time s RRI ms i5 HeartRate 1 59 1 7 5 i 5 teat oc 1 4 ME NN NN NS 10 15 Time HeartRate T Click the Edit Plot icon a A plot menu will open With the help of the plot menu it is possible to modify a lot of plot parameters For example you can adapt the figure to a special layout wanted for a special publication Please see also chapter General Software Features 57 Cardio2D Manual Click average Ave tab in single channel view ED Monitor 2 0 45 1 40 40 2 44 Rec 3 45 2 3l Rec 4 DL 20 10 0 10 250 300 350 400 Time ma Q QR Oo 9 The recordings of one user defined electrode channel are displayed in a color coded overlay plot Each recording represents for example one concentration indicated by a different color The number of the recording is displayed beside
18. with intracellular electrodes and field potentials with extracellular electrodes have shown that there is a linear relationship between the rise time of the cardiac action potential AP and field potential FP as well as between AP and FP duration The contribution of different ionic transmembrane currents can be identified in the shape of the FP waveform as well as shown in the following picture The correlation between the waveform components and the ion channel activities was shown by using ion channel blockers or depleting the medium of the respective ions You can clearly see the rapid component of the depolarizing sodium current and the slow calcium current 2000 1000 e FP uv 1000 2000 uie ee wee ene pee 1 88 1 83 Time s1 The delayed rectifying current IKr is represented either by a positive or negative peak The polarity of the peak depends on several parameters for example the proximity of the cell layer to the measuring electrode and cannot be predicted which does not matter for this assay The field potential duration corresponds to the action potential duration which can be correlated to a QT like Interval in an electrocardiogram It is measured from minimum of the Na peak to the maximum minimum of the IKr current peak 116 Electrocardiogram PR Segment a QRS Complex U I I I I I I I I
19. Apply lt 1016 13 gt Recording 1 Test lt 10 16 19 gt Apply lt 10 16 21 gt Recording 2 Test lt 10 16 26 gt Apply lt 10 16 30 gt Recording 3 Test lt 10 16 55 gt Apply lt 10 16 36 gt Recording 4 Test lt 10 16 42 gt Apply lt 10 16 44 gt Recording 5 Test lt 10 16 49 gt Apply lt 10 17 08 gt Recording Test lt 10 10 09 gt Esperment stop 34 Cardio 2D Software 4 7 Stimulation of MEA Electrodes Click Stimulate icon stimulate Device MCS S T 40072 Channel mulus Syne Symmetric 1 Phase SOO ps Extension 300 us 2 Phase 500 ps IPI 99 mz The stimulation feature gives the possibility for voltage driven stimulation of a MEA electrode This is useful for example if a cardiomyocyte culture did not form a pacemaker This dialog allows to connect a stimulus generator STG and to design the stimulation pulse You can connect either one stimulus generator STG with more than one channels or several STGs to stimulate the different MEA amplifiers Define either the channel number from the Channel drop down menu when using one STG or the devices from the Device drop down menu when using several STGs EE Device MLS STG4002 20033 Channel MCS STG4002 20041 35 Cardio2D Manual Stimulus Pulse In the Stimulus menu you design the stimulus pulse Stimulus Symmetric yl 1 Phase 2 Phase IPI
20. Click the Start icon The progress of analyzing in dependency of the different recording phases is displayed Recording 114 79 Cardio2D Manual Click HB Average tab in Analyze dialog da analyze X HB Detection 20 On Recording 3 Fre HB Interval ET ms Fast HB Interval FOO ms me Apply to all Recordings Recording If the heartbeat average feature is welcome please enable the check box On If you do not use this feature do not enable the check box to save computer performance Select the Recording for the heartbeat average from the drop down menu Define the time interval before a heartbeat Pre HB Interval and the interval after a heartbeat Post HB Interval from the up down boxes During these time spans no additional heartbeat should be detected Make sure to keep these intervals shorter than the shortest heartbeat to heartbeat interval Click the button Apply to all Recordings and the settings are applied to all available recordings phases Otherwise you can define different settings for each single recording phase If different values for different recordings are set this will be indicated by the orange background color of the drop down box Please see the next picture lw Recording Pre HB Interval ms Post HB Interval ms Apply to all Recordings 41 80 Theoretical Background Click 2D tab in Analyze dialog
21. Detection 56 HB detection setate io Vic orae 56 56 LabDDOOK iuis na E 56 Cardio2D Manual 56 56 MEA Types 10 120 200 30 10 256 9wellMEA 10 60 6wellMEA 10 10 60MEA200130 10 40 1000 10 60MEA500 10 10 GOMEAS5OO 30 10 ii iod rave DOS 10 a uds 10 2 10 MEA Window 56 MEA Window Cardio2D 56 MEA Window Cardio2D 92 MEA2100 10 N Noise level 71 Offset Correction 17 P pacemaker 63 Q QT related Analysis 112 Dose 112 QT like Interval 112 R ROI Selection 100 Recording Sequence 100 Remove 100 S Settings File 89 Single channel window 17 56 Software 7 7 7 124 Data Acquisition 7 System requirements 7 USB ME32 FAl System
22. The white menu bar Building a Stimulus Pattern in the middle of the dialog is for creating the desired stimulus pattern from the provided default stimulus patterns called Primitives or for the import of external created stimulus patterns You can modulate each Primitive pattern via individual Parameter Settings and the modulation will be immediately displayed in the Single Pattern view and also in the view for the Complete Stimulus Pattern 41 Cardio2D Manual Building Stimulation Patterns On the left side of the upper section you can choose the provided default stimulus pattern Move the primitive icon via drag and drop into the white field in the middle of the dialog for creating a user defined stimulation pattern Add as much primitives as needed It is possible to change the sequence of the primitives by drag and drop For deleting one of the pattern please drop it into the bin Set Marker Signal Additionally it is possible to set a marker signal Open the Set Marker Signal dialog by clicking J Marker the check box Set Marker 77 in the window for parameter settings Set Marker Signal X Marker Offset ps 60 Duration ps 30 Click the check box Set Marker and click Repeat if you want to repeat the marker signal at each cycle of the stimulation pattern Adjust the Offset in us and the Duration in us of the marker signal via the up down boxes The or
23. are build analogical On the left side of the dialog you can choose the part of recording you are interested in from the drop down menu Select Recording above the MEA field Select the designated electrode channel on the MEA electrode field On the right side the analysis parameters provided for export are listed Define the parameters you like to export via check box and create a file by pressing the Export button Export File j 20120323 01 1 RecO E21 Heartbeats csv Notepad File Edit Format View Help Maximum praesta us Maximum Amplitude nv Minimum Timestamp us Minimum Amplitude nv Peak to Peak Amplitude nv Peak to Peak Duration us Peak to Peak Slope v s Max Slope v s Max Slope Timestamp ys Max Slope Amplitude cut abet Rate Hz Inter Beat Interval us 1495300 1910265 1495600 2118298 4028563 300 13 429 25 787 1495450 389285 2496500 1901677 2496800 2126886 4028563 300 13 429 25 773 2496700 1044911 0 999 1001200 3528300 1903485 3528600 2118298 4021783 300 13 406 25 796 3528500 1006491 0 969 1031800 4676050 1923599 4676350 2115586 4039185 300 13 464 25 911 4676200 417648 0 871 1147750 5831250 1915576 5831550 2108580 4024156 300 13 414 26 415 5831400 612573 0 866 1155200 7011300 1931509 7011600 2143610 4075119 300 13 584 24 944 7011500 1250345 0 847 1180050 8271000 1927328 8271300 2139994 4067322 300 13 558 25 052 8271150
24. are different the respective input field will be highlighted in orange 29 Cardio2D Manual For example Type the desired settings into the Study tabbed page of the global labbook selected by the icon in the Control View Confirm the entry with Enter During writing the background of the input field will be orange after confirming the window is white again Close the global Labbook Settings dialog Open the local labbook via icon in the MEA window The local settings will be changed The other way round when changing settings locally differing values will be highlighted in orange in both dialogs This feature comparing global and local settings is also available for settings in the Analyzer Settings dialog Labbook Settings xi 1 5 study General study General FileName MEAID FileName 1 Comment Comment Comment Comment Click General tab in Labbook Settings dialog WSLabbook Settings EF MEA 1 Labbook Settings x Study General Study General Department Department Department Department In this dialog you fill in general data referring to your experiment 30 Cardio 2D Software 4 6 Compound Basic Compound Click Compound in main menu Experiment The Basic icon is replaced the Compound icon Click the Compound icon n The Compound Sett
25. beating rate of all heartbeats of one recording phase Select the number of the displayed MEA electrode in the Selection control window of the Control window on the left side 97 Cardio2D Manual Single channel view and the color map of the Heartbeat tab correspond with each other The excitation pattern of that heartbeat marked with a blue arrow is exactly shown in the color map Selecting a new heartbeat with the blue arrow the color map will be adapted immediately The user has four possibilities to analyze the heartbeats Click near to any heartbeat and it will be marked with the blue arrow This way you can jump to any heartbeat of that recording phase Or you can analyze each single heartbeat and the related color map step by step by clicking the icons Previous K and Next Ei Or you can run through all heartbeats automatically with Play gt and stop the performance with Stop fa Or you can click onto a data point in the plot of the Latency Velocity window MEA Electrode 4000 2000 FP uv c 2000 0 59 0 592 0 594 0 596 0 595 Time s 10 ms In the MEA electrode window the field potential of exactly that electrode is displayed which is selected in the color map via mouse To adjust the size of the displayed raw data interval please use the up down control box Click Edit Plot icon 3k A plot menu appears With the help of the plot menu it is possible to modify a lot of plot pa
26. channel in the heartbeat window of the Control Window on the left side of the HB Averager tab The channels are color coded the legends respectively and the number of heartbeats averaged is stated in brackets To adapt the y axis please use the right drop down menu You can display the recording or apply phases only or all phases Please use the left drop down menu 106 Theoretical Background If you like to zoom the Natrium peak of the heartbeats for QT like analysis please click the icon Na Peak w From the same recordings displayed before now the Natrium peak only is automatically displayed in this window 200 41 02 18 3 tL LL 100 200 Time ma Testonly 100 2 Channel Overlay 109 21 31 22 30 50 23 3 E 51 30 52 30 TE 50 TU 0 200 400 Time m 100p v ep In the Channel Overlay window the heartbeats of the different recording phases of one single channel are displayed in a overlay plot The recording phases of the channel are color coded and the number of heartbeats averaged in this recording is stated in brackets To adapt the y axis please use the drop down menu 107 Cardio2D Manual 6 8 Analysis of Field Potential Duration Field Potential ROI Selection Heartbeat HE OT Analysis QT Interval ms Dose Response 100 1000 10000 100000 Dose ph Absolute QT related Interva
27. da analyze Cluster Channels Max Propagation Time 20 ms Recording o The first up down box in this tab of the dialog defines the minimal number of cluster channels The Min Cluster Channels gives the minimum number of channels at which heartbeats have to be detected to form a cluster For example for a MEA electrode layout with 60 electrodes it could be 10 but if less than 60 electrodes are involved it could be 5 or 2 The second up down box defines the maximum propagation time of the heartbeat within a cluster for example the maximum duration between the first and the following detection of a heartbeat on different electrodes 81 Cardio2D Manual Click QT tab in Analyze dialog analyze X HB Detection HB Average 2D QT On Detection Deadtime Recording o This dialog defines the parameters for the QT related Analysis QT related Analysis is possible only if the heartbeat average is enabled QT related Analysis will always be performed on the interval from the heartbeat time stamp to the last data point of the average heartbeat The parameter Detection Deadtime defines a time window after the heartbeat time stamp within which no analysis will be performed to avoid detections on artefacts originating from the sodium peak 82 Theoretical Background Help B Cardio2D File Tools CardiazD4 Help Abaut CardiazD4 Astemizale Menu for activa
28. the Windows operating system interfere with the data storage on the hard disk and can lead to severe performance limits in Cardio2D These routines were designed for use on office computers but are not very useful for a data acquisition computer Deselect Windows Indexing Service for all local disks Turn off Optimize hard disk when idle automatic disk fragmentation It is also not recommended to run any applications in the background when using Cardio2D Remove all applications from the Autostart folder Important Please do not run MC Rack software program from Multi Channel Systems MCS GmbH in parallel to Cardio2D software Please check the system requirements before you install the software MCS cannot guarantee that the software works properly if these requirements are not fulfilled Important Please make sure that you have full control over your computer as an administrator Otherwise it is possible that the installed software does not work properly Cardio2D Manual Installing the Cardio2D Software Make sure to have a valid license for the Cardio2D software Please ask license multichannelsystems com You do not need a license for the Cardio2D software if you like to test the software for getting an idea of how it works and what is possible You can test all functions run the viability test to see your data but you are not able to start an experiment Please contact license multichannelsystems com to receive a valid licen
29. the mean value of four or five points to calculate the interpolated point Show Electrodes Click the check box for displaying the black circles indicating the electrodes in the color map JE Click the Map Settings icon The dialog Map Settings appears Click Data tab Field Potential Map Settings Data Map Interpalatian Missing Data Nearest eighbor Missing Channel Show Time Stamp hhmmss hhmmss ms hhmmss In the Map tab of the Map Settings dialog the interpolation parameter refers to the color of the map In the Data tab the interpolation parameter refers to the data generating the map Default is the interpolation method Nearest Neighbor Please see above With the method of interpolation you can substitute missing channels also The program uses the same Nearest Neighbor method for interpolation If you do not want to substitute missing channels click Off In the drop down menu Show Timestamp you can choose the type of the timestamp you want to be displayed below the color map 93 Cardio2D Manual Click the Record Movie icon The dialog Movie Recorder appears MEA 1 Movie Recorder a Frame Hate 30 Frames s Compression On E File Mame Stark Cancel The movie of the color map will be recorded in AVI format The AVI format is a common format You just need a Media Player to replay the video You can choose the Fram
30. the plot and the number of averaged segments are denoted in the brackets behind bonto 3 45 Aec 40 Rec 44 3 Please specify the length of the averaged segments Analyzer Settings HB Average tab The data in Monitor are displayed but not saved in the data file Please read also chapter Basic in Control Window 4 16 Latency Velocity Window Latency 5600 us Monitor 0 1 Rec2 Rec 3 Ree 4 Latency ms Time s e QQlRo Mla dlou Latency 58 Cardio 2D Software The diagram of Latency window shows the latency us of the excitation calculated between the first and the last detected electrode as a function of the time s whereupon the time is the duration of the experiment from starting the recording The actual value of the latency is displayed above the plot in red letters The different recording phases are displayed color coded in overlay propagation path The latency is added up from point to point that means the result is not zero when defining a circular path Please start a viability test and define the propagation path Do not change the path later during the experiment is running Please read also chapter Propagation Path in Color Map Click Edit Plot icon 3k A plot menu appears Poo B amp Alk With the help of the plot menu it is possib
31. the signal dependent amplitude the waveform detection is more flexible The detection routine detects amplitudes in relationship to the noise level The amplitude is calculated with its standard deviation in a range from 1 to 50 It is recommended to use a signal dependent amplitude The standard deviation is used to estimate the individual amplitude change for each channel separately A time interval of 1 s is used to calculate the standard deviation You set the factor by which the standard deviation is multiplied 1 to 50 To compensate individual differences in the noise level of channels it is recommended to use the signal dependent amplitude as detection parameter The data is scanned for a situation where the minimum amplitude either fixed or relative to the standard deviation is met and the rise time is in the defined range as well Limit the rise time range to exclude artifact signals that are in the range of the defined amplitude As the sodium peak is generally very fast small rise times such as 100 to 400 us can usually be used The higher you define the FP change parameter the higher you need to set the rise time as well as a higher FP change needs longer than a smaller one The rise time and the amplitude of the Na Peak are used to detect only signals that show the expected kinetics of the waveform Slow potential shifts are excluded Automated Waveform Detection Min Amplitude Min Ris
32. together with a 256 9wellMEA and each of the nine wells is displayed and controlled separate in nine MEA window tabs MEA A to MEA MEA 1 MEA 2 MEA 3 MEA 4 ue If you connect for example an USB ME256 System for data acquisition it is possible to connect up to four MEAs MEA_1 to MEA_4 Each MEA window tab will display the heartbeats with different settings in the four views of the MEA Tabbed Pages in the MEA Window independent from each other Via the Control Window you start and stop the experimental protocol simultaneously 19 Cardio2D Manual Control Window The experimental protocol is visualized by three icons on the left orange control view The icons can be clicked for editing the protocol Below the control view the log file documents the ongoing experimental proceedings Summary Window The Summary window is only available if more than one MEA is connected It presents an overview about the layout of the MEAs which are connected and you can select or deselect the recording of connected MEAs The summary corresponds with the MEA tabbed pages in the MEA Window MEA Window The MEA Window is divided into four single views All of them display heartbeats in different characteristics If more than one MEA is connected to the experimental setup each MEA can be operated separately via the MEA Tabbed Pages Depending on the hardware up to nine MEAs for nine wells of the 256 9wellMEA are available Se
33. 0 10 MEA 3 MEA 658 100 10 MEA 4 MEA 100 10 Close Data Acquisition with the MC_Card The data acquisition computer with the with the internal data acquisition board MC_Card comes preinstalled and preconfigured by MCS for a flawless operation You should contact your local retailer for assistance if you want to install additional hard or software or if you want to replace the computer as incompatibilities of hardware components or software settings with the MC Card or Cardio2D may occur ES Data Acquisition Settings X Device 743 Sample Hate T Hange 4096 mV Resolution Channel Diver Version 1444222 MEA 1 MEA 8 8 100 10 Software and Hardware Installation Data Acquisition with the USB MEA256 Systems The USB MEA256 System is a compact and portable stand alone solution for MEA recordings with integrated amplification data acquisition and analog digital conversion The system acquires data from up to 252 electrode channels four additional analog channels and 16 digital IN OUT channels The digitized electrode data is transmitted to the connected computer via universal serial bus High Speed USB 2 0 Thus it is possible to use any computer as a data acquisition computer also a laptop Recorded data is graphed analyzed and reviewed with Cardio2D and Cardio2D program The integrated filter amplifier supports 252 electrode channels The gain of 1100 is fixed The ban
34. 0 30 MEA 9Well 256MEA 300 30 USB ME32 EcoFlexMEA 6x6 300 50 FlexMEA 6x6 300 30 16 Software and Hardware Installation USB MEG64 MEA 6Well 60MEA 200 30 MEA 8x8 100 10 MEA 8x8 200 30 MEA 8x8 1000 30 MEA 6x10 500 10 MEA 6x10 500 30 8x8 700 100 Qwane 60 6x10 400 70 Qwane 60 6x10 500 75 Qwane 60 6x10 600 80 FlexMEA72 750 100 FlexMEA 6x6 300 30 Note It is possible to connect one or two FlexMEAs EcoFlexMEA 6x6 300 50 Note It is possible to connect one or two EcoFlexMEAs USB ME 128 MEA 6Well 60MEA 200 30 MEA 8x8 100 10 MEA 8x8 200 30 MEA 8x8 1000 30 MEA 6x10 500 10 MEA 6x10 500 30 8x8 700 100 Qwane MEA60 6x10 400 70 Qwane MEA60 6x10 500 75 Qwane MEA60 6x10 600 80 FlexMEA72 750 100 FlexMEA 6x6 300 30 Note It is possible to connect one or two FlexMEAs per 68 pin MCS standard connector EcoFlexMEA 6x6 300 50 Note It is possible to connect one or two EcoFlexMEAs per 68 pin MCS standard connector EcoFlexMEA 12x12 2700 100 17 Cardio2D Manual USB ME256 MEA 6Well 60MEA 200 30 MEA 8x8 100 10 MEA 8x8 200 30 MEA 8x8 1000 30 MEA 6x10 500 10 MEA 6x10 500 30 8x8 700 100 Qwane MEA60 6x10 400 70 Qwane MEA60 6x10 500 75 Qwane MEA60 6x10 600 80 FlexMEA72 750 10 FlexMEA 6x6 300 30 Note It is possible to connect one or two FlexMEAs per 68 pin MCS standard connector EcoFlexMEA 6x6 300 50 Note It is possible to connect one or two EcoF
35. 2D Analyzer Program OVerVIelWW ce ucc uda 75 Cardio2D Control window 75 89 Cardio2D File information 75 Cardio2D Filed potential tab 75 92 Cardio2D Heartbeat 75 101 Cardio2D Heartbeat tab 75 101 Cardio2D QT related Analysis 112 Cardio2D ROI Selection 100 Cardio2D Color map 75 92 101 Cardio2D File 75 Cardio2D Help 75 Cardio2D Overlay plots 75 92 Cardio2D Re analyzing 75 Cardio2D Single channel view 75 92 101 Cardio2D Tools 75 1 75 Color map window 17 63 COH TUR 63 Da 63 Interpolation of 63 Isochrone color 63 ISOCHFOPGS YER 63 Map Settings iii big 63 interpolation of color 63 SIZE AD Usate 63 Record 63 COMPTESSION pr TEC 63 Frame rate 2 UT e Rd 63 Movie format AVl 63 Compound 31 Contact Information 121 Control Window 17 27 IE 17 27 COMMEN 27 Set MarKear xvid etes PH 27 27 VADY use pt uo ut te ttp 17 27 Running a viability test
36. Data tab the interpolation parameter refers to the data generating the map Default is the interpolation method Nearest Neighbor Please see above With the method of interpolation you can substitute missing channels also The program uses the same Nearest Neighbor method for interpolation If you do not want to substitute missing channels click Off In the drop down menu Show Timestamp you can choose the type of the timestamp you want to be displayed below the color map Open the dialog in menu Tools Analyze Raw Data d CardiozD File Tools Help Analvze Raw Data Theoretical Background Click 2D tab in Analyzer Settings da analyze X HB Detection HB Average 20 HT Cluster Channels LJ Propagation Time 20 mz T Recording o The setting for the Propagation Path is from minimum to maximum by default If you like to define a propagation path manually for example when propagation is not linear but circular please select the option Manual Path This option enables the option Set Propagation Path in the color map Latency Velocity Select Latency from the drop down menu Recording 0 amp Recording 1 4 TOVE Recording 8 The diagram shows the latency ms of the excitation in dependency of the time ms between the minimum and the maximum color coded in overlay for each recording separately 10 Latency ms 30 Time
37. GmbH has been advised of the possibility of such damages 3 1 Software and Hardware Installation Installing the Cardio2D Software System Requirements Software One of the following Microsoft Windows operating systems is required XP Vista Windows 8 1 or Windows 7 English and German versions supported with the NT file system Other language versions may lead to software errors Hardware Please read chapter Hardware Acquisition Systems One of the listed data acquisition hardware system MEA2100 System USB ME Systems USB MEA256 System USB ME32 FAI Systems or MC Card is recommended by MCS Due to the amount of recorded data a computer with low performance may lead to performance problems therefore Multi Channel Systems recommends an up to date computer Please contact MCS or your local retailer for more information on recommended computer hardware specification Please note that there are sometimes hardware incompatibilities of the data acquisition system and computer components or that an inappropriate computer power supply may lead to artefact signals Important You need to have installed the latest data acquisition driver to operate the device which is automatically installed with Cardio2D The installation may be invalid if the data acquisition does not respond Please contact Multi Channel Systems MCS GmbH or your local retailer in this case Recommended Operating System Settings The following automatic services of
38. MEA 8x8 100 10 MEA 8x8 200 30 MEA 8x8 1000 30 MEA 6x10 500 10 MEA 6x10 500 30 8x8 700 100 Qwane MEA60 6x10 400 70 Qwane MEA60 6x10 500 75 Qwane 60 6x10 600 80 14 Software and Hardware Installation MC Card 64 e MEA 6Well 60MEA 200 30 e MEA 8x8 100 10 e MEA 8x8 200 30 e MEA 8x8 1000 30 MEA 6x10 500 10 e MEA 6x10 500 30 8x8 700 100 Qwane 60 6x10 400 70 Qwane 60 6x10 500 75 Qwane 60 6x10 600 80 FlexMEA72 750 100 FlexMEA 6x6 300 30 Note It is possible to connect or two FlexMEAs EcoFlexMEA 6x6 300 50 Note It is possible to connect one or two EcoFlexMEAs MC Card 128 MEA 6Well 60MEA 200 30 MEA 8x8 100 10 e MEA 8x8 200 30 MEA 8x8 1000 30 e MEA 6x10 500 10 e MEA 6x10 500 30 8x8 700 100 e Qwane 60 6x10 400 70 Qwane 60 6x10 500 75 Qwane 60 6x10 600 80 FlexMEA72 750 100 e FlexMEA 6x6 300 30 Note It is possible to connect one or two FlexMEAs per 68 pin MCS standard connector e EcoFlexMEA 6x6 300 50 Note It is possible to connect one or two EcoFlexMEAs per 68 pin MCS standard connector 15 Cardio2D Manual USB MEAG4 MEA 6Well 60MEA 200 30 MEA 8x8 100 10 MEA 8x8 200 30 MEA 8x8 1000 30 MEA 6x10 500 10 MEA 6x10 500 30 8x8 700 100 Qwane 60 6x10 400 70 Qwane 60 6x10 500 75 Qwane 60 6x10 600 80 USB MEA256 MEA 16x16 10
39. Max Search 20 ms HB Timestamp Minimum Recording o In this dialog you define the parameter for detecting the waveform of a field potential Please see also chapter Automated Waveform Detection Because you have to define these settings before recording data the same dialog appears in Cardio2D MEA electrode window Analyzer Settings The first parameter for detection is the Rise Time Select the minimum and the maximum of the rise time from the drop down menus The range involves 100 us to 2 ms for the minimum rise time and 200 us to 5 ms for the maximum rise time Next parameter is the Amplitude You can decide to have a fixed amplitude for detection or a signal dependent amplitude The fixed amplitude involves a voltage range from 10 up to 1 mV Using the signal dependent amplitude the waveform detection is more flexible The detection routine detects the amplitude in relationship to the background noise The amplitude is calculated with its standard deviation in a range from 1 to 50 It is recommended to use a signal dependent amplitude The following parameters characterize the detected field potential Detection Deadtime defines the time span in between no further field potential should be detected The detection dead time has to be longer than the time used for searching the minimum or maximum otherwise the following error message appears SSS xl The Detection Dead Time has be longer tha
40. a acquisition device if more than one data acquisition is available Select the Sample Rate the Range when using the MC Card only and the gain as an intrinsic property of the connected amplifier The Resolution the number of Channels and the Driver Version are displayed Select the type of MEA or FlexMEA Please save these entries with the command Save Defaults Otherwise the dialog will appear each time you want to start Cardio2D It is possible to change the default settings or to save several default files if you work with different configuration The dialog is also available in the Settings menu in Data Acquisition You will be asked for a path to put the log and data files The program will create subfolders for each of these file types It is advisable to put all three subfolders in a common Cardio2D directory These settings can be changed later please see main menu Settings File Location Location Settings Haw Data Dokumente und Einstellungen sCardia2D ata Results C Dokumente und Einstellungen LardiazD D ata ER Lag Files C Dokumente und Einstellungen sCardia2D D ata Close In case you have to reinstall the software you will be asked for the file locations again Note It is recommended to save the settings in the Data Acquisition Settings dialog as default This way you do not have to type them in again when restarting Cardio2D 3 3 Cardio2D Manual For saving the
41. ab provides parameters which indicate the effect of different doses of a compound on 2D signal propagation across the MEA 113 7 1 7 2 Cardio2D Manual Troubleshooting Troubleshooting Most problems occur seldom and only under specific circumstances In most cases it is only a minor problem that can be easily avoided or solved If the problem persists please contact your local retailer The highly qualified staff will be glad to help you Please inform your local retailer as well if other problems that are not mentioned in this documentation occur even if you have solved the problem on your own This helps other users and it helps MCS to optimize the instrument and the documentation Please pay attention to the safety and service information chapter Important Safety Advice Multi Channel Systems MCS GmbH has put all effort into making the product fully stable and reliable but like all high performance products it has to be handled with care Error Messages NENNEN CardiazD is recording data Stop measurement before program exit Please stop recording before exit the Cardio2D program CardiazD DataldeFaulEMEA 1 20 already exists Override Please enter new file with new file name Be aware that Ja will overwrite present data Choose Nein to enter a dialog and save data in a new file Error in Series string Please check the syntax Message Convert Dose incorrect
42. ack number on the connected computer The hardware configuration microprocessor frequency main memory hard disk of the connected computer This information is especially important if you have modified the computer or installed new hard or software recently Test Model Probe The provided test model probe simulates a MEA with a resistor of 220 kiloohms and a 1 nanofarad capacitor between bath and electrode for all 60 electrodes and can be used for testing the MEA amplifier Test Model Probe GND bath Pad electrode 119 9 3 Cardio2D Manual Contact Information Local retailer Please see the list of official MCS distributors on the MCS web site User forum The Multi Channel Systems MCS GmbH User Forum provides the opportunity for you to exchange your experience or thoughts with other users worldwide Mailing list If you have subscribed to the Newsletter you will be automatically informed about new software releases upcoming events and other news on the product line You can subscribe to the list on the contact form of the MCS web site www multichannelsystems com 120 10 Index 121 Cardio2D Manual 122 Automated Waveform Detection 3 B Basic 31 Blanking Circuit BC 36 Blanking 36 EXTENSION 36 SVNC QUE RM tus EDU 36 36 Cardio2D 1 Cardio2D Program Overview 17 Cardio2D Analyzer 1 Cardio
43. amp can be set to minimum or maximum or maximum slope in the HB Timestamp drop down menu in the Analyzer dialog Please click on the Analyzer icon in the MEA Electrodes plate and enter the HB Detection tab in Analyzer Settings The red line labeled with the identifier of the selected channel indicates that the zoomed MEA channel was switched Click Edit Plot icon 3k A plot menu appears ee With the help of the plot menu it is possible to modify a lot of plot parameters For example you can adapt the figure to a special layout wanted for a publication Please see also chapter General Software Features 56 Cardio 2D Software Click heartbeat HB tab in single channel view The HB tab shows a diagram of the duration of the RR interval in ms if you select RRI ms from the drop down menu In whole heart experiments the RR interval is defining one complete heartbeat from one ventricular depolarization to the next ventricular depolarization 1 RR is the beating frequency of the heart Any modulation of the beating frequency is an indicator for possible arrhythmia So arrhythmia or proarhythmic events can be detected In cell cultures ECG terminology does not apply However the depolarization driven by rapid sodium channels causes an as distinctive peak as the QRS complex in the ECG Analysis can be performed in strict analogy to the ECG analysis in respect of signal propagation patterns and conduction velocity
44. ange colored marker signal will be displayed in the window for the Complete Stimulus Pattern 42 Cardio 2D Software Modulation of the provided Stimulation Patterns Initially the provided stimulation patterns are default To adjust a primitive to your requirements please click onto the icon which will be highlighted in pale blue In the upper part of the dialog the setting parameters for this pattern appears on the left and a picture of the actual shape of the pattern on the right Immediately after changing a pattern the modulation is visible in the Single Pattern view and in the Complete Stimulus Pattern view Save the stimulus pattern as default by clicking the Save Primitive as Default icon I1 beside the Marker check box Most of the parameters have to be adjusted with up down boxes Please click into the up down box and move the wheel of your mouse for quick adjustments in wide steps Use the arrows for fine tuning The modulation is immediately displayed in the single pattern and in the complete pattern window Overwrite the letter in the up down box the modulation will be displayed after confirming the value with Enter or after clicking into another box Playing with the different possibilities for adjustment allows to create each shape of pulse which could be necessary Additionally it is possible to modulate the shape when repeating pulses in relationship to each other Stimulus Pattern Flat Line m The first p
45. anged in a grid according to the MEA layout The channel identifier refers to the columns and rows of the MEA layout grid The drop down menus at the bottom of the window are for scaling the axes MEA 1 MEA 2 ju TU LLL LLL ete BEBO HOURLY 200 uv ENS The electrodes number 15 and 32 without field potentials are grounded The electrode number 56 highlighted in dark green is the electrode which is displayed in the Single channel view Please read the next chapter 55 Cardio2D Manual 4 15 Single Channel View Click field potential FP tab in single channel view Ch 7B Ch 55 58 54 2000 1000 0 2000 2000 l c 73 74 75 76 77 78 79 8n 8 82 Time s 500 105 iu The single channel view allows a closer view to one of the MEA electrode channels You can zoom to any channel by clicking on a channel in the MEA electrodes plate The zoomed channel is displayed in the larger single channel view The plot shows the field potential against the time s To scale the axes please use the drop down menus The range for the field potential is 1 uV to 5000 pV the time can vary between 1 and 60 seconds Reset the axes by clicking on the Reset icon 7 The detected heartbeats are marked with black dots The detection timest
46. anual Slope Calculation The calculation of the slope of each data point P is achieved by linear regression This linear regression is performed within a time interval around each data point Increasing the interval will help to reduce noise on the slope The time interval around a data point P can be defined by the user in the Analyze dialog of the QT related Analysis tab in Cardio2D Please read also chapter QT related Analysis Analyze X HB Detection HB Averager 20 HT k Detection D eadtime 50 ms E Slope Calculation 15 0 ma Select time span for the Slope Calculation in ms in the up down box to define the size of the time interval which is used for slope calculation of each point of the signal Please see the picture ms P Interval for slope calculation 70 6 1 Cardio 2D Analyzer Software The Cardio2D Analyzer Program Overview File Tools Help Field Potential ROI Selection Heartbeat HB Averager QT Analysis AstemizoleMEA_1 c2d AstemizoleMEA 1 c2r LUE UE HHHIHHHHHHHH H HH EHE H E single Channel View 39 5 3 Channel View Recording Control Control fa El m Ei s m Cardio2D Results Labbook Environment Experiment Stimulate DataSource DeviceBus DeviceN ame VendorlD ProductlD 10892900 Color Map Setting File Veg ut Wem zx
47. asure the membrane potential that results from the electrochemical gradient directly with an intracellular electrode When ion channels are opened due to chemical or electrical stimulation the corresponding ions are moving along their electrochemical gradient In other words the resistance of the membrane is lowered resulting in an inward or outward flow of ions measured as a transmembrane current The extracellular space is conductive as well and though the resistance is very low it is not zero According to Ohm s law lt the extracellular current results in a small voltage that can be measured with extracellular electrodes Extracellular signals are smaller than transmembrane potentials depending on the distance of the signal source to the electrode Extracellular signal amplitudes decrease with increasing distance of the signal source to the electrode Therefore a close interface between electrode and cell membrane is very important for a high signal to noise ratio The transmembrane current and the extracellular field potential follow the same time course The field potential is roughly equal to the first negative derivative of the transmembrane potential thus revealing an information that is comparable to conventional methods This has been shown for different types of signals derived from neuronal preparations as well as cardiac preparations This convenient method is used by Cardio2D Simultaneous recordings of action potentials
48. ately after installation The test model probe is already mounted on the amplifier It simulates a MEA with a resistor of 220 and a 1 nF capacitor between bath and electrode Use the Cardio2D program to record from the test model probe and to check the amplifier This also presents an opportunity for you to learn about the program before starting a real experiment Please keep the test model probe at a safe place in case that you will need it later for troubleshooting Starting the recording For testing the amplifier it is not necessary to run a complete experiment You may choose to do so to learn to know the program On the Control panel click the Viability icon Data is not recorded You see the raw data streams of all 60 channels in the typical MEA layout grid You may have to adjust the position and span of the axes until you can clearly see the noise level Click any channel to zoom into this channel Following screen shot shows the typical behaviour of a MEA amplifier with test model probe 67 Cardio2D Manual 5 2 Detection Parameter EF Analyzer Settings HB Detection HE Averager 20 Minimum Rize Time 200 us Maximum Aize Time FOO us C Fixed Amplitude Signal Dependent 10 StdDev Detection Deadtime 200 ms Min Max Global x Min Mas Search 10 ms HB Timestamp Minimum Under the tab HB Detection in Analyzer Settings of the MEA electrodes plate of the Card
49. con 7 The dialog Movie Recorder appears MEA 1 Movie Recorder Movie Type Te Frame Hate 30 Frames s Compression File Mame Stark Cancel The movie of the color map will be recorded in AVI format The AVI format is a common format You just need a Media Player to replay the video You can choose the Frame Rate between one and hundred frames per second Select Compression of the movie with On or deselect the compression with Off Save the movie by clicking the Save icon al and select a file name and the path to the folder you want to store it Recording the color map movie starts as soon as you click Start 63 Cardio2D Manual 4 18 General Software Features Tracking Axes RR RE nF og 53 61 65 M E p u a Qik o i ale id 500gv 10s i p For tracking the axis manually click on the axis and the mouse pointer will change into a hand mid To scroll the axes please move the mouse with the hand pointer For zooming the axes press the CTRL key and keep it pressed while moving the mouse or the wheel of the mouse The mouse pointer turns into a double arrow symbol A right mouse click on the axes gives four options Edit 500 Tracking Enabled Update Resume values Zoom Fit In View Edit Tracking Enabled Update Rescue Values an
50. ct So the appropriate time has to be tested empirical depending on the experiment Click the Set Propagation Path icon in the color map window After selecting the option in the color map you can define a propagation path manually by clicking on the electrodes of the color map For deselecting electrodes of the path please click the electrode again 00 00 00 589650 The setting for the Propagation Path is from minimum to maximum by default If you like to define a propagation path manually for example when propagation is not linear but circular please click the icon Set Propagation Path in the color map After selecting the option in the color map you can define a propagation path manually by clicking on the electrodes of the color map For deselecting electrodes of the path please click the electrode again 99 Cardio2D Manual When changing the propagation path from the minimum maximum default to manual both displays in the Latency Velocity window are immediately re calculated and adapted 2200 us a 5342100 ps The electrodes on the MEA are symbolized by circles Black circles indicate a detection of a heartbeat on this electrode in the currently displayed cluster white circles indicate no detection of a heartbeat in the currently displayed cluster but the color map is completed by interpolation of data JE Click the Map Settings icon tab Propagation Map Settings T
51. ct So the appropriate time has to be tested empirical depending on the experiment The Min Cluster Channels gives the minimum number of channels at which heartbeats have to be detected to form a cluster For example if a syncytium covers all 60 electrodes of a MEA it could be 10 otherwise it should be less than 10 51 4 10 4 11 Cardio2D Manual Start Stop Start Stop Click Start to start recording The Start icon changes into Stop icon Sto and the duration of recording is counted up in seconds Stop recording manually by clicking on Stop icon Cardio2D generates three types of files the Raw Data file c2d the Result file c2r and the Settings file c2s Raw data file and the associated settings file always belong together in the result file the settings file is included The log file is not automatically generated but by users command Log File Below the control window is the log file window Each step of your experiment is documented together with its timestamp You can save the log file by clicking on the Save icon The log file has the extension xml Under Settings in File Location you define the path for the log file To clear the log file display please click the Clear icon X The message Cleared Log 15 22 46 gt Cleared Lag will appear lt 06 44 20 gt Cardio2D start lt 09 44 21 gt Add MEA 1 lt 09 07 17 gt Viability start lt 09 09 24
52. d Zoom to Fit With the command Tracking enabled you can move the axis by moving the mouse or by moving the mouse wheel in horizontal direction on the x axis and in vertical direction on the y axis Edit Click Edit and the Plot Axis Editor appears You can adjust general and specific parameters for your plot layout Plot Axis Editor x General Title Display Scale Range Grid Lines Tracking Dock 1 Visible Title T ext Time s E Enabled Context Menu Enabled Color AM Layer 10 o User Can Edit Min 46 339146495623 Can Focus 5 261 45550424 76 Cantrol Key Toggle Enabled Span 20923507 0084953 Master LII Scale Type Linear Reverse To Cursor Scaler 64 Click the Plot icon The following toolbar appears amp ois m Tracking Scroll axes Zoom axes Zoom in and zoom out Select Zoom box Data and Cursor Edit Copy to clipboard Save Print Preview Cardio 2D Software Resume All Resume Selecked Picture Copy Data 65 Cardio2D Manual Click Edit icon w The following dialog appears E Conto write d E Control xm riv Now it is possible to format the plot 66 5 1 Experimental Proceeding General Performance Noise Level Please use the provided model test probe to test the amplifier immedi
53. de rn 15 20 Duration lt 130 ES 440 100 Marker 45 Cardio2D Manual Stimulus Pattern Ramp Modulate the amplitude Amplitude mV and the duration Duration us of the ramp pulse via up down boxes To adjust the steepness of the arms of the pulse please use the three up down boxes for the ascending part the plateau and the descend arm independent from each other Setting the number of cycles to more than one enables the inter stimulus interval ISI ps up down box Additionally the Arrow button will be enabled Amplitude mV 100 Duration lt 1000 1000 1000 E Cycles 1 at 151 us E When using more than one cycle you can click onto the arrow symbol for additional options A highlighted parameter field appears to modulate the amplitude and the duration of the ramp pulse in relationship to each other The window Single Pattern shows the ramp pulse in an overlay plot The lower window Complete Stimulation Pattern shows the combined sequence of all stimulus pulses Amplitude m 5 Duration lt 4810 E 2060 200 z Cycles 11 a ISI us 3200 Amplitude rn 5 Duration lt 40 ES 40 Marker 46 Cardio 2D Software Stimulus Pattern Biological Pulse Import Ampl 5 15 ps i 0 0 If you like to use for example a biological signal as stimulus pulse you can
54. dwidth of 1 Hz to 3 kHz is suitable for a broad range of applications Data Acquisition with the USB ME32 FAI Systems The USB ME32 FAI System is a very compact and portable stand alone solution It features an integrated 32 channel filter amplifier and data acquisition The digitally converted electrode signals are transmitted to the connected computer via universal serial bus High Speed USB 2 0 As in other USB ME Systems it is possible to use any computer as a data acquisition computer also a laptop The size of the complete system is small enough to fit into a laptop bag together with the computer The system is for use with a 32 channel miniature amplifier MPA32I You can directly connect the EcoFlexMEA36 to the 32 channel miniature preamplifier for recording with Cardio2D Overview Possible Configuration of Data Acquisition System and Microelectrode Array MEA MEA2100 MEA2100 HS 120 MEA 12x12 200 30 MEA2100 HS2x60 MEA 6Well 60MEA 200 30 MEA 8x8 100 10 MEA 8x8 200 30 MEA 8x8 1000 30 MEA 6x10 500 10 MEA 6x10 500 30 8x8 700 100 Qwane 60 6x10 400 70 Qwane MEA60 6x10 500 75 Qwane 60 6x10 600 80 13 Cardio2D Manual IMEA2100 HS60 MEA 6Well 60MEA 200 30 MEA 8x8 100 10 MEA 8x8 200 30 MEA 8x8 1000 30 MEA 6x10 500 10 MEA 6x10 500 30 8x8 700 100 Qwane 60 6x10 400 70 Qwane 60 6x10 500 75 Qwane 60 6x10 600 80 MEA2100 Lite HS60 MEA 6Well 60MEA 200 30
55. e 4 lt 07 33 07 gt Viability start lt 07 33 43 gt Viability stop lt 07 34 15 gt Viability start lt 07 34 30 gt Viability stop 24 You uncouple the Summary window to move it to a different place the monitor Click the je Map Settings icon to open the Summary Map Settings dialog and start or stop monitoring the color map via Start and Stop icon Summary Map Settings Data Color Map Jet Invert Contour 1 ms Maximum Auto Size HJ 200 200 Interpolation Bicubic Show Electrodes vl Close 25 Cardio2D Manual In this example the Summary page shows 60 6wellMEAs with adapted color map Summary MEAE 2 MEAS Iv 4 Help ES rardin2D File Experiment Tools Settings CardiazD Help About Cardio2D License Kev Menu for starting the Cardio2D online help to monitor the dialog About Cardio2D and to enter the license key For a valid license please read chapter Installing Cardio2D Software Select About Cardio2D About Cardio2D L ardia2D Version 2 2 1 0 Channel Systems MES GmbH Copyright MES 2007 2013 Close The dialog shows basic information about the version of Cardio2D Note Please keep in mind that the information of this dialog is necessary in ca
56. e MEA window on the right side is divided into separate window panes and has five tabs Field Potential ROI Selection Heartbeat HB Averager and QT related Analysis Switching between the tabs influences the settings of each of the separate windows Select the single channels of each tab via Selection control window of the Control Window of Cardio2D 86 Theoretical Background 6 4 Field Potential Cardio2D Click Field Potential tab Field Potential ROI Selection Heartbeat HB Averager Analysis Single Channel View MEA Electrode Field i Li m O Channel Overlay BK H H gt The Field Potential tab is divided in four separate windows The Single channel view the MEA electrode field the Channel overlay and the Color map Single Channel View The single channel view displays the field potentials of one recording phase of the MEA electrode selected in the Channel control With the Zoom Box icon 1 4 you can zoom to any region of the data trace Click the Reset Axes icon 10 reset the default settings of the axes 87 Cardio2D Manual Click Edit Plot icon 3k A plot menu appears 54 With the help of the plot menu it is possible to modify a lot of plot parameters For example you can adapt the figure to a special layout wanted for a special publication Please see a
57. e Rate between one and hundred frames per second Select Compression of the movie with On or deselect the compression with Off Save the movie by clicking the Save icon al and select a file name and the path to the folder you want to store it For recording the color map movie please click Start and press Play in the Channel overlay window 94 Theoretical Background 6 5 ROI Selection Click ROI Selection tab Field Potential Selection Heartbeat Average HT Analysis 100 Analyze 0 D 1 2 3 4 5 it 8 Time s 4 Analyze 01 D 1 2 3 4 5 8 9 Time s 1 02 TEUER mer Eee em m e PN 0 1 2 3 B 4 1 Recording Sequence 05 0 1 d 3 4 5 4 1 11 Analyze 04 100 100 uw Time s 95 Cardio2D Manual The ROI Selection tabbed page lists the single recording sequences from one electrode Select the electrode in the Channel control field of the Control Window Via ROI you can discard one part or several parts of the recording sequence which are now excluded from the analysis Additionally it is possible to discard a complete recording sequence by deselecting the respective check box The data in the ROI will not be included in the analysis 100 R
58. e Time Max Rise Time 2 08 2 0 8 wd 2 8 mef This illustration shows how the Cardio2D routine detects the fast sodium peak by the three parameters field potential change minimum and maximum rise time The software scans the recorded data points and it checks whether following data points meet the required criteria For example when the waveform detection routine arrives at the data point marked with the red line in the figure the following data points meet the amplitude criterion shaded in gray The signal reaches this field potential change in the preset range of the rise time shaded in green The data point at which both the FP change and the rise time criteria are met is marked with a red dot in this figure Would the signal be smaller than the minimum FP change or faster or slower it would not be detected In other words the signal needs to cross the bottom border of the green shaded area Any signal component that matches the criteria will be detected The FP change parameter is the main parameter that should discriminate biological signals from noise The rise time parameter ensures that biological or artefact signals that meet the FP change criterion but have a shorter or longer rise time than the sodium peak are not detected Please read the next chapter HB Detection in The Cardio2D Analyzer Program Overview about detection parameters in Cardio2D 69 Cardio2D M
59. e icon Stop the recording manually whenever it is reasonable Icon The recording time span is user defined Rec 4 Rec 0 Rec Rec 2 Rec 3 gt In these types of recording described above the data of each recording interval is completely saved in the recording file To avoid a big amount of data it is additionally possible to save the data of a recording only while the Apply Phases between the recordings are displayed but excluded from the data file This may be the phase during experimental manipulation for example while pipetting new compounds and waiting for the effect Enable the check box Apply Phase Start the recording manually The first phase of this experiment is the Apply phase and the dialog Set Marker show up Write a comment if necessary and manipulate the experiment To finish the Apply Phase and to start the recording phase please click the button Set Marker Comment Hifedipin The data of the recording phase is saved the data of the apply phase is not saved in the recording file The data of the apply phase is displayed in the single channel view marked by the term Monitor Moritor 3 45 Hec 1 40 44 Hec 3 33 Cardio2D Manual Apply Phase The recording time is user defined The data of the recording phase is saved the data of the apply phase is excluded from the data file Apply Rec Apply Rec Apply The log file on the left s
60. ec 02 0 1 2 3 4 5 8 4 1 Time 5 To set a Region of Interest ROI please click with the mouse into the desired recording sequence and draw a red box by keeping the left mouse button pressed To change the breadth of a ROI use the red buttons on the left and on the right side to replace a complete ROI click into the red field and move it with the mouse MN Remove Marker To delete the ROI click with the right mouse button into the red box The command Remove Marker will appear Or click onto the ROI to select it and press the delete key 500 uv Select an value in uV from the drop down box to scale the axes of all recording sequences Use the buttons for zooming in and resetting the axes for each recording phase separately 96 Theoretical Background 6 6 Heartbeat Cardio2D Click Heartbeat tab Field Potential Selection Heartbeat Average HT Analysis 22 Single Channel View I cc m Em CIE 33 124 33 125 33 128 zala 33 132 SI 9500 us MEA Electrode zi Recording 00 00 33 1 23600 Latency ms Color Map BIT eu 3l 34 Latency Velocity Latency The Field Potential tab is divided in four separate windows The Single channel view the MEA electrode the Latency and the Color map Single Channel View Field Potential Heartbeat The single channel view displays the amplitude and the
61. ecording 0D 00 33 1 23500 Latency ms Color Map cbe eu 3l 34 Latency Velocity Latency 19 Select HB Averager tab Please see the picture below Important Please pay attention to the Analyze dialog To use the average tool you have to enable the check box in the HB Average tab of the Analyze dialog in main menu Tools da analyze X HB Detection HBAverage 20 aT k Recording Fre HB Interval 50 ms ES rn Fast HB Interval ms ES rs Apply to all Recordings 74 Theoretical Background The MEA electrode field window corresponds directly with the Channel overlay view The field potential of one selected channel or of several selected channels are shown in overlay in the Channel overlay window Select the channel s via mouse click in the MEA electrode field Select one channel from the Heartbeat window pane of the Control Window and the desired channel will be displayed with its different recordings in the Recording Overlay window You can choose whether you want to display all phases or test or apply phase only from the drop down menu Field Potential All Selection Heartbeat HB amp verager HT Analysis 01 41 21 33 Rec 05 41 31 33 05 20 41 33 62 33 51 33 Recording Overlay Channel Overlay 100 L
62. eld Potential tab Please see the upper picture The single channel view on top of the right window panes shows field potentials Recorded data traces are displayed in the MEA electrodes pane If you select one or more than one MEA electrodes in this window pane by clicking on the desired channel they are displayed in the overlay plot window below The color map displays the spreading of the excitation of a heartbeat from the pacemaker or the earliest time point detected if the pacemaker is not on the MEA to the outer regions of the cardiac cell culture or tissue The isochrones are indicated by black lines The colors indicate the time distance between the first signal and outer regions of excitation Red color represents the pacemaker when using the color map jet inverse Below the map the actual timestamp is displayed The color map corresponds with the single channel view and with the field potential windows 72 Theoretical Background Select ROI Selection tab Please see the picture below Field Potential Selection Heartbeat HB Average HT Analysis 100 Analyze 00 D 1 2 3 4 5 8 9 10 Time s Analyze 01 D 1 2 3 4 5 8 9 10 0 Time s 1 02 ae eese eoi eren pecie e
63. eld potential It does not matter whether the Sync Out check box in the stimulation dialog is selected or not it will have no effect Amplifier with Blanking Circuit BC The blanking feature is used to reduce stimulus artefacts in MEA recordings A TTL pulse the blanking signal that has to be timed exactly with the stimulus pulse triggers both the stimulation and the blanking During the blanking signal both stimulus inputs are connected to the software selected stimulation electrode s while all recording electrodes are disconnected from the amplifier The last signal value on all electrode channels before the blanking pulse will be saved and then kept until the channels are reconnected to the amplifier This avoids that the amplifier gets saturated on all channels during stimulation which is the case with amplifiers without blanking circuit 37 Cardio2D Manual MEA Select Select 15 a software program for controlling the blanking circuit of the MEA amplifier With MEA Select it is possible to select the MEA electrode you want to stimulate Start MEA Select parallel to Cardio2D The MEA Select main window appears Select Version 1 2 0 x Stimulation 7 Not Stim 47 Ground Blanking Configuration Load Save o Ww ail Help 100 Lis Values are rounded to multiples of 20 ps 21 a1 a si en 1 12 22 32 e se 2 72 ee is fea sia ue
64. gt Viability stop lt 09 09 29 gt Experiment start lt 19 09 26 gt Recording Test lt 09 10 11 gt Esperment stop lt 09 10 30 gt Experiment start lt 09 10 30 gt Recording 0 Test lt 09 10 56 gt Recording 1 Test lt 09 10 41 gt Recording 2 Test lt 09 10 47 gt Recording 3 Test lt 09 10 55 gt Recording 4 Test lt 09 10 58 gt Recording 5 Test lt 09 11 01 gt Esperment stop lt 09 11 05 gt Esperment start lt 09 11 05 gt Recording Test lt 09 11 11 gt Recording 1 Test lt 09 11 16 gt Recording 2 Test lt 09 11 22 gt Recording 3 Test lt 09 11 22 gt Expernment stop 52 Cardio 2D Software 4 12 Summary Window Fils Expenrent Took d usu Latency 3900 ps Summary Window Click View Summary Page in main menu Settings to open or to remove the Summary window This Summary window is only available if more than one MEA at a time is connected The window presents an overview and all connected MEAs are visible The dark green color code spotlights the MEA which is displayed in the MEA tabbed page The layout of the MEA is displayed respectively The screenshot for the example above was taken with a MEA2100 2x60 System This system has two slots for MEAs with 60 electrodes When using 60 6wellMEAs each of the MEA has six separate chambers with nine electrodes and a reference electrode in each well The layout of the MEA is disp
65. he dialog Propagation Map Settings appears Click Data Map Color Map Jet k Invert Contour Off 1 ms Arrays 3 ms Auto Size wi x 290 x 290 Interpalatian Bicubic Show Electrodes yl In this dialog you have to define the settings for the color map 100 Theoretical Background Color Map The usual used color map for cardiac issues is the Jet Invers map but you can also choose Hot Cool or Grey maps To remove the colors from the map click Off The isochrones will remain only Contour The drop down menu is for changing the color of the contour for the isochrones into Black Jet Hot Cool and Grey Or you can remove the lines with Off Adjust the number of the contour lines from the numeric box beside up to 100 contour lines are possible Arrows The feature is not yet implemented in this version of Cardio2D As soon as it will be implemented you find a Read Me on the MCS web site where you will be able to download the new version of Cardio2D Minimum Maximum You have to define the minimum and the maximum of the scale of the color map That means you define the time frame in which the excitation pattern are displayed in the color map Usually you start with zero in the minimum that is the pacemaker The excitation pattern will run from the pacemaker over the map to the maximum The widest selectable ti
66. he reference is not integrated in the plot In Absolute plots of the dose response curve the standard reference of 0 uM is set to 1 uM and displayed in the logarithmic scale Data Export to ASCII Click the Plot icon a The following toolbar appears Please read also chapter General Software Features 4 55 The data displayed in the different windows can be exported to ASCII format by clicking a the icon Save 110 6 9 Theoretical Background Export of Results To be free for further analysis of the results from Cardio2D the software offers finitely many possibilities to export results to ASCII format The export file is organized as a comma separated value file csv Please open the Result Export dialog via main menu File option Export B Cardio2D Open Raw Data Open Result File The Result Export dialog opens Heartbeat Heartbeat 2D Averaged Heartbeat Dose Response Dose Response 2D Na Peak Select Recording Recording 0 no dose specified Maximum Minimum Peak to Peak Amplitude in Peak4o Peak Duration Peak4o Peak Slope Max Slope Heart Rate Frequency Inter Beat Interval 8 8 298 8 2 8 82 Export Heart Beats The dialog has five tabs Heartbeat Heartbeat 2D Averaged Heartbeat Dose Response Dose Response 2D The tabs of the analyzer tools
67. ide of the window shows following entries lt 10 16 19 gt start lt 10 15 19 gt Recording 0 Apply lt 10 15 20 gt Recording 1 Test lt 10 15 26 gt Recording 2 Apply lt 10 15 27 gt Recording 3 Test lt 10 15 32 gt Recording 4 Apply lt 10 18 36 gt Recording 5 Test lt 10 15 42 gt Recording Apply lt 10 16 42 gt Recording Test lt 10 15 48 gt Recording 8 Apply lt 10 15 50 gt Recording 9 Test lt 10 16 55 gt Ewperment stop Enable the check box Apply Phase and the check box Record Start the recording manually The first phase of this experiment is the Apply phase and the dialog Set Marker show up Write a comment if necessary and manipulate the experiment To finish the Apply Phase and to start the recording phase please click the button Set Marker The data of the recording phase and the data of the apply phase is saved in the recording file Additionally the different types of data in the file are marked as data from the apply or the recording phase Apply Phase and Record The recording time is user defined The data of the recording phase and the data of the apply phase is saved in the data file Rec Rec 5 Rec Rec 3 Apply Appl Appl The log file on the left side of the window shows following entries lt 70 16 02 gt start lt 10 16 05 gt Apply lt 10 16 04 gt Recording 0 Test lt 10 16 10 gt
68. import signals by clicking the Import button The imported file must mandatory have the following format 3479512 dat Datei Bearbeiten Format Ansicht 45147 300 19072 500 21903 700 0 800 11026 1000 21307 1200 18178 1500 21158 1600 9387 1800 17880 4 14 Timestamp Voltage Value Timestamp Voltage Value Timestamp Voltage Value The unit for the timestamp is us and for the voltage value nV The units are not part of the file Cardio2D software accepts integers only and commas tabulators or spaces to separate the timestamp and the voltage value Please remove a possible header and use a new line for each integer pair Save the file with the extension dat Adapt the amplitude of the signal in percent 96 via up down box Setting the number of cycles to more than one enables the inter stimulus interval ISI us up down box Additionally the Arrow button will be enabled 100 000 Cycles 50 000 151 us 0 50 000 100 000 Ampl 2 zi 1 000 000 Marker Cardio2D Manual When using more than one cycle you can click onto the arrow symbol ti for one additional option A highlighted parameter field appears to modulate the Amplitude 96 in relationship to each circle of the signal The window Single Pattern shows the imported pulse as overlay plot The lower window Complete Stimulation Pattern shows the combined sequence of all stimulus pulses Tool Bar
69. ings dialog appears E Compound Settings X Series 0 ph 100 1 pM 10 100 phi Switch Recording Timer Test Duratian s Record Apply Phase 4 Close In this dialog you can choose series of compounds which are applied to the cardiac cells or tissue on the MEA In parallel to the Basic Settings dialog the options referring to the recording phases are available Please see above There are two series of compounds provided The dilution factor in Series 1 10 is ten and in Series 1 3 the factor three But you can create a user defined compound series by overwriting the given example in the window Please consider the syntax otherwise you get a syntax error message Error in Series string Please check the syntax Message Convert Dose incorrect parameter The functionality of the settings in the Switch Recording drop down menu is the same as in the Basic settings Define the duration of the test phase in the Test Duration drop down menu Enable the check box Record Apply Phase if you want to save all data inclusive the data of the apply phase in the recording file 31 Cardio2D Manual In the control window beneath the Compound icon the currently applied dose of compound is displayed When clicking on the Compound icon the Set Marker dialog appears which is called here Apply Compound dialog in the header the next dose to apply is additionally displayed Type your c
70. io2D program you can adjust the parameters for the field potential detection Sometimes it will be necessary to play with this parameters until you find the best configuration for your experiment Analyzer Settings xl Settings X HB Detection HE amp verager HB Detection HB Averager 20 On Propagation Time 10 me Pre HE Intewdl 50 ms Min Cluster Channels 9 Post HB Interval Click Analyze Raw Data in menu Tools in Cardio2D program The same dialog will appear as in the HB Detection tab in Analyzer Settings of the MEA electrodes plate of the Cardio2D program please see above In the dialog Analyze you can adjust the parameters for the field potential detection as well 68 5 3 Experimental Proceeding The sodium current is detected by the rapid change in the field potential indicated by the rise time The rise time refers to the time required for a signal to change from a specified low value to a specified high value Typically these values are 10 and 90 of the step height The output signal of a system is characterized also by fall time If the rise time is chosen very short the slope of the peak is very steep if the rise time is longer the slope will be smooth You have to try out which time is the best to characterize the Na Peak you want to detect You can either define a fixed value for the amplitude of the field potential for example 100 Using
71. l 200 400 600 444 ms 474 ms The Analysis of Field Potential Duration is also called QT related Interval The field potential duration corresponds to the action potential duration which can be correlated to a QT like Interval in an electrocardiogram It is measured from minimum of the Na peak to the maximum minimum of the IKr current peak The QT related Interval tabbed page is until now divided in two separate windows Please select an appropriate channel from the heartbeat window of the Control Window On the left side the QT related Interval window the different recording phases of the selected channel are displayed on the right side a Dose Response curve is shown if possible to calculate Important Please pay attention to the Analyze dialog To use the QT related Analysis tool you have to enable the check box in the HB Averager tab of the Analyze dialog in main menu Tools and the respective check box in the QT related Analysis tab 108 Theoretical Background da analyze X HB Detection HB Averager 2D HT On Detection D eadtime 50 ms Slope Calculation 15 0 ma Enable the check box On to use the QT related Analysis tool Select the Detection Deadtime in ms in the first up down box to define the time span in which no detection is performed Detecting extremal values for QT related Analysis is achieved by analyzing the slope of the signal Select the Slope Calc
72. layed in the summary page the well which is highlighted in dark green appears in the MEA window tabbed page and can be operated via control window Deselect the check box of a well to exclude the electrodes from recording Click the Map Setting icon je to adapt the color maps in the summary page Click Pause Monitoring icon to stop displaying the color map to save performance capacity Summary 4 Dockable 1 MDI Child The summary page is dockable to different places on monitor Click the icon Window Position z and decide where to place the summary window In Floating mode you can push the window wherever you want It is also possible to drag and drop each window into the desired position Click the Auto Hide icon and the summary page will be hidden Right Top Bottom 53 Cardio2D Manual When clicking on the Summary button the window will be visible again m MEA Electrodes m Color Map met tr H MEA window Monitor 2 Rec 45 Rec1 140 2 44 Rec3 27 Velocity mmms 0 200 400 Single Channel View Latency Velocity 50017 The MEA Window is divided into four window panes all showing heartbeats in different characteristics The settings made in MEA Window influence all four smaller windows respectively The prominent MEA Elec
73. le to modify a lot of plot parameters For example you can adapt the figure to a special layout wanted for a publication Please see also chapter General Software Features Change the latency window via drop down menu to velocity window Velocity 0 27 mm ms Monitor 0 1 2 3 4 9 gt mmm Hg The diagram of Velocity window shows the velocity mm ms of the excitation calculated between the maximum and the minimum of excitation as a function of the time ms whereupon the time is the duration of the experiment from starting the recording The actual value of the velocity is displayed above the plot in red letters The different recording phases are displayed color coded in overlay propagation path The velocity is added up from point to point that means the result is not zero when defining a circular path Please start a viability test and define the propagation path Do not change the path later during the experiment is running Please read also chapter Propagation Path in Color Map 59 Cardio2D Manual 4 17 Color Map The color map displays the propagation of each heartbeat across the MEA The isochrones are indicated by black lines in this screenshot but the user can change their color in Contour The colors map is represented as false color Below the map the actual timestam
74. lect the respective MEA in the Summary window to display the data traces in the MEA window If a MEA is disabled via check box in the Summary window the corresponding MEA tabbed page will be removed from the display and no data is recorded Different types of MEAs are implemented The standard MEA with 60 electrodes and 8 x 8 electrode layout grid the diameter of the MEA electrodes and the spacing between them differs 60MEAs with 6x10 electrode layout grid the 120MEA with 12 x 12 layout grid the 256MEA with a 16 x 16 layout grid the 256 9wellMEA with nine separate wells shown in the picture above or the EcoFlexMEA36 with 6 x 6 electrode layout grid The design of the MEA window is automatically adjusted to the type of the MEA please see chapter Data Acquisition Settings in First Use of Cardio2D after Installation Recorded data traces are displayed in the MEA electrodes pane You can zoom to any channel by clicking on a channel in the MEA electrodes plate The zoomed channel is displayed in the larger single channel view The single channel view has three tabs FP field potential HB heartbeat and Ave average The heartbeat window shows either a diagram of the RR interval or of the frequency of the heartbeats the heart rate The current values of the RRI or the frequency are displayed above in red numbers The color map displays the propagation of a heartbeat from the pacemaker to the outer regions of the cardiac cell cultu
75. lexMEAs per 68 pin MCS standard connector EcoFlexMEA 12x12 2700 100 18 4 Cardio 2D Software 4 1 The Cardio2D Program Overview ini x File Experiment Tools Settings Help T MEA Electrodes Color Map E LJ Control Window lt 11 42 07 gt Apply 100 uM lt 11 42 25 gt Recording 4 Test 100 ph lt 11 42 59 gt Experiment stop Monitor 1 45 Rec 1 40 Rec2 44 ly 100 nM 1 Rec3 45 32 4 Recording Test 100 nN 2 b Rec 4 Rec Rec2 Latency ms Log File amp PU puer lt 13 24 58 gt Recording 4 Test 100 ph lt 13 26 03 gt Experiment stop 3 1 Single Channel View Main Window Overview The main window of Cardio2D consists of two in case of several connected MEAs of three window panes The left Control Window is for controlling and documentation of the experiment The bigger window on the right side the MEA Window displays heartbeats in different characteristics in real time monitoring The Summary window is only available if more than one MEA is connected You have one single Control Window only but depending on the connected hardware you can have up to nine different MEA tabbed pages in the MEA Window which correspond with the Summary window respectively If you connect for example an USB MEA256 System for data acquisition you can use it
76. ling the software Click Save Defaults to save your settings as a default This way your setting parameters will be present each time you start Cardio2D To restore default settings please select Restore Defaults Experiment FA CardiozD File Experiment Se Basic Compound Menu to schedule an experiment Selecting Basic or Compound changes the icons in the Control Window Basic defines the default settings for an usual experiment Compound gives the possibility to apply an user defined series of compounds 21 Cardio2D Manual Tools ES rardio2D Experiment Tools Settings Help Offset Correction Menu to activate the offset correction if necessary Enable the option Offset Correction and the following dialog appears WS Offset Correction x Corect Offset An offset correction may be necessary if you observe disturbing voltage offset on the input channels Click the check box Correct Offset and Cardio2D takes 100 ms of the recorded data in the moment when the check box is selected to calculate the DC offset The mean of this 100 ms sweep is subtracted from the recorded data as long as the Correct Offset check box is activated The offset correction works separately for each available channel Click the Update button to update the calculation of the offset correction This option is available when Cardio2D is running Settings ES rardio2D Experiment Tools
77. list knowledge and training and have received instruction in the use of the device have read and understood the chapter on safety and the warning instructions in this manual and confirmed this with their signature It must be monitored at regular intervals that the operating personnel are working safely Personnel still undergoing training may only work on the device under the supervision of an experienced person 2 4 2 5 Important Information and Instructions Terms of Use for the Program You are free to use the program for its intended purpose You agree that you will not decompile reverse engineer or otherwise attempt to discover the source code of the software Limitation of Liability Multi Channel Systems MCS GmbH makes no guaranty as to the accuracy of any and all tests and data generated by the use the software It is up to the user to use good laboratory practice to establish the validity of his findings To the maximum extent permitted by applicable law in no event shall Multi Channel Systems MCS GmbH or its suppliers be liable for any special incidental indirect or consequential damages whatsoever including without limitation injuries damages for data loss loss of business profits business interruption loss of business information or any other pecuniary loss arising out of the use of or inability to use the program or the provision of or failure to provide Support Services even if Multi Channel Systems MCS
78. lso chapter General Software Features The two red bars in the heartbeat plot of the single channel view represent the region of interest ROI Click on the middle of one red bar near to the zero line The mouse pointer will turn into an arrow with two arrowheads Keep the mouse button pressed Now you are able to vary the breadth of the ROI The displays in the MEA electrodes window and in the overlay plot window are adapted immediately Additional you can click anywhere on one of the red bars and the mouse pointer will turn into a cross with four arrowheads Keep the mouse button pressed Now you can move the ROI with constant breadth to any region you want to analyze closer Again the displays in the MEA electrodes window and in the overlay plot window are adapted immediately Click with the right mouse button on one of the red bars The Edit menu appears Data iew Editor EB X General Fil Inside Fil Outside Dock Name Data View 1 Visible v Enabled Context Menu Enabled Title Text Dataview 1 E LserCanEdt Can Focus Color Laver 100 2H arid Linies Mirror Dr ataView Hame Vertical Horizontal Axes Control Enabled Mouse Style Mone kd Wheel Style Keyboard Style Both m Theoretical Background MEA Electrode Field Recorded data traces are displayed in the MEA electrode field To adapt the y axis please use the drop down menu The MEA electr
79. me frame for the color map spans from 0 us in the minimum to 10000 ms in the maximum To change the colors of the map please use the first drop down menu Color Map Auto Click the check box Auto in Maximum option The Minimum is zero you are not allowed to modify it in Cardio2D The Cardio2D program automatically finds for the maximum of each heartbeat and adapts the scaling of the color map It is recommended to use the auto scaling function Size W x H You can modify the size of the color map Interpolation Interpolation is a mathematic method to calculate missing information between two points of a line The bilinear interpolation uses four points of the nearest vicinity to calculate the interpolated point The bicubic method uses eight points to calculate the interpolated point The interpolation method of the nearest neighbor uses the mean value of four or five points to calculate the interpolated point Show Electrodes Click the check box for displaying the black circles indicating the electrodes in the color map 101 Cardio2D Manual Click the Map Settings icon M The dialog Propagation Map Settings appears Click Data tab Field Potential Map Settings Data Map Interpalatian Missing Data MearestMeighbor Missing Channel Off Show Time Stamp hhmmss hhmmss ms hhmmss In the Map tab of the Map Settings dialog the interpolation parameter refers to the color of the map In the
80. multichannel 4 systems Cardio2D Manual Information in this document is subject to change without notice No part of this document may be reproduced or transmitted without the express written permission of Multi Channel Systems MCS GmbH While every precaution has been taken in the preparation of this document the publisher and the author assume no responsibility for errors or omissions or for damages resulting from the use of information contained in this document or from the use of programs and source code that may accompany it In no event shall the publisher and the author be liable for any loss of profit or any other commercial damage caused or alleged to have been caused directly or indirectly by this document 2015 Multi Channel Systems MCS GmbH All rights reserved Printed 18 11 2015 Multi Channel Systems MCS GmbH AspenhaustraBe 21 72770 Reutlingen Germany Fon 49 71 21 90 92 5 0 Fax 49 71 21 90 92 5 11 info multichannelsystems com www multichannelsystems com Microsoft and Windows are registered trademarks of Microsoft Corporation Products that are referred to in this document may be either trademarks and or registered trademarks of their respective holders and should be noted as such The publisher and the author make no claim to these trademark 1 1 1 2 2 1 2 2 2 3 2 4 2 5 3 1 3 2 3 3 4 1 4 2 4 3 4 4 4 5 4 6 4 7 4 8 4 9 4 10 4 11 4 12 4 13 4 14 4 15 4 16 4 17 4 18
81. n a new dialog enabling the definition of the stimulus pattern as explained on the following pages To start stimulation with the currently in the second section of the tool bar x selected pattern only click the Start icon Third section To clear all sites click the Clear all stimulus sites icon Stimulation of electrodes produces stimulation artefacts which can be eliminated To select the Start all stimulus patterns simultaneously via the Start icon in this section blanking artefact suppression please click the Blanking icon ue Blanking will blank all recording electrodes on a MEA while a stimulus pattern is applied to any electrode of the same MEA Choosing the Dedicated Electrode icon i all selected electrodes will be defined to be dedicated stimulation electrodes These electrodes will be permanently isolated from the amplifier hence recording from these electrodes will be stopped Note It is not possible to select electrode 15 for stimulation because this electrode is connected to the internal reference Viability Test Before you start recording a viability test is recommended The data is not recorded During viability test it is possible to change the stimulation electrodes and stimulation pattern without stopping the display of the data Such changes are not possible during data acquisition You first have to set stimulation electrodes and stimulation parameters which are fix and then star
82. n define the settings for the color map 61 Cardio2D Manual Color Map The usual used color map for cardiac issues is the Jet Invers map but you can also choose Hot Cool or Grey maps To remove the colors from the map click Off The isochrones will remain only Contour The drop down menu is for changing the color of the contour for the isochrones into Black Jet Hot Cool and Grey Or you can remove the lines with Off The time step is defined in the dialog Arrows The feature is not yet implemented in this version of Cardio2D As soon as it will be implemented you find a Read Me on the MCS web site where you will be able to download the new version of Cardio2D Minimum Maximum You have to define the minimum and the maximum of the scale of the color map That means you define the time frame in which the excitation pattern are displayed in the color map Usually you start with zero in the minimum The excitation pattern will run from the pacemaker over the map to the maximum The widest selectable time frame for the color map spans from 0 us in the minimum to 10000 ms in the maximum Auto The Cardio2D program automatically finds the minimum and maximum of each heartbeat and adapts the scaling of the color map It is recommended to use the auto scaling function Size W x H You can modify the size of the color map Interpolation Interpolation is a mathematic method to calculate missing information in between
83. n the Min Max Search Interval 78 Theoretical Background The drop down menu Min Max Type describes the type of the minimum or maximum Local or Global These options refer to one single field potential signal Local means the first detected minimum or maximum global means the deepest minimum or highest maximum in the field potential Please see the picture 2000 1000 FP 1000 Local Global 2000 minimum minimum DE i LEM HS IM IESU MEC QUI GEN 1 88 1 83 Time s Whether you select the local or the global minimum or maximum depends on the aim of the user If you are interested in the maximal slope of the action potential you have to select the first local minimum or maximum If you are interested in absolute minimal or maximal values you have to select the global minimum or maximum Please select the time used for searching the minimum or maximum of the field potential Min Max Search from the drop down menu range from 1 ms up to 200 ms You can set a HB timestamp to each detected field potential either to the maximum or to the minimum or to the maximal slope of the field potential This timestamp is visualized in the plot with a black dot and stored in the raw data file Please read also chapter Detection Parameter Re analyzing If you change in any tab of the Analyze dialog parameters for the analyzed file it is possible to re analyze the data with the new settings
84. nd the software are not intended for medical uses and must not be used on humans Malfunctions which could impair safety should be rectified immediately High Voltage Electrical cords must be properly laid and installed The length and quality of the cords must be in accordance with local provisions Only qualified technicians may work on the electrical system It is essential that the accident prevention regulations and those of the employers liability associations are observed Each time before starting up make sure that the mains supply agrees with the specifications of the product Check the power cord for damage each time the site is changed Damaged power cords should be replaced immediately and may never be reused Check the leads for damage Damaged leads should be replaced immediately and may never be reused Do not try to insert anything sharp or metallic into the vents or the case Liquids may cause short circuits or other damage Keep the device and the power cords always dry Do not handle it with wet hands Requirements for the installation Make sure that the device is not exposed to direct sunlight Do not place anything on top of the device and do not place it on top of another heat producing device Never cover the vents not even partially so that the air can circulate freely Otherwise the device may overheat 2 2 2 3 Cardio2D Manual Guarantee and Liability The General conditions of sale and delive
85. ode field corresponds with the overlay plot window The data traces of the MEA electrodes selected in the MEA electrode field and highlighted in white are displayed in the overlay plot window in different colors EE EEEIEE A A A E 200 uv The MEA electrode field refers also to the single channel view In single channel view you define the region of interest Data of this ROI frame are shown in each channel of the MEA electrode field and in the overlay plot window If the ROI will be modified in the single channel view data traces in MEA electrode field and in overlay plot window are adapted immediately 89 Cardio2D Manual Channel Overlay If you select one or more than one MEA electrodes in the MEA electrode field they are displayed in an overlay plot in the Channel overlay window To adjust the y axis please use the drop down menu There is a prominent red bar in the diagram The function is different compared to the two red bars in single channel view limiting the ROI This red bar here defines the timestamp belonging to the currently shown excitation patterns of the color map Please expand the x axis to have a closer view to the displayed field potentials Click the icon Snapshot 6 to move the red bar timestamp step by step please click the icons Step Backward and Step Forward The color map will be immediately adapted To create a movie of the excitation pat
86. omments in the Comment window Apply Compound 100 pM X Apply Compound 1 nM Comment Comment apply to all MEAs apply to all MEAs Basic Click Basic in main menu Experiment The Compound icon is replaced by the Basic icon The recordings are counted up and displayed WS Basic Settings Task Phase Switch Duration Count Apply Phase Apply Phase Hecord The Basic Settings dialog is for scheduling the recordings 32 Cardio 2D Software There are different ways to influence the recordings Select Timer in the Switch drop down menu and define the phase duration 1 s to 30 min from the Duration drop down menu Start the recording manually A new recording phase will start automatically after the predefined recording interval is finished Stop the recording manually whenever it is reasonable Timer The recording time span is fixed 305 305 305 305 30 s 305 Rec 0 Rec 1 Rec 2 Rec 3 Rec 4 Rec 5 Select Timer and define a Duration of a recording phase Enable the check box Count and select the number of recordings in the up down box Start the recording manually The recording will stop automatically after the selected number of repeats Select Click Icon in the Switch drop down menu Now you define the recording time manually by clicking onto the icon Start the recording manually A new recording phase will start immediately after clicking th
87. ossible to display several recordings of an electrode channel in overlay and to average the values In this dialog the pre and post time before and after a heartbeat can be selected via up down box Enable the check box On to display and save the averaged data traces If you do not need the average function disable the check box to save computer performance and disk space 50 Cardio 2D Software Click 2D tab in Analyzer Settings EF Analyzer Settings ES Mea 1 Analyzer Settings X HB Detection HB Averager 20 HB Detection HB Averager 20 Propagation Time 10 me Propagation Time Him H Cluster Channels 5 Cluster Channels In this dialog you can define the time span from the drop down menu range of 1 200 ms in which the Propagation Time should be calculated The action potential runs over the heart cell culture or tissue or whole heart from the pacemaker to the peripheral regions of the syncytium The excitation propagation is measured in high spatial resolution by the MEA electrodes detecting cardiac field potentials and extracting local activation times The propagation time is the longest possible period for passing of the excitation from the first detected excited electrode to the last detected excited electrode of one cluster If the propagation time is too short you miss data for the complete heartbeat If the propagation time is too long the next heartbeat starts and the results are incorre
88. ou can check the signal detection parameters and focus onto single channels If the settings are too rigid the cardiac signal is not detected at all Too loose settings will lead to a wrong signal detection and thus to wrong results After adjusting all parameters in the different window panes you can start recording The log file documents the experimental proceeding It is possible to change analysis parameter during viability test is running to check the differences immediately This option is not available during recording the settings have to be constant during the whole time of recording Labbook Click Labbook icon E The dialog Labbook Settings will open Click Study tab In this dialog you fill in data referring to your experiment The Labbook Settings will be stored in the settings file 25 Comparing Local and Global Settings If more than one MEA is connected to the experimental setup or a multi well MEA you can control each MEA or each well separately via local settings or you can control all MEAs together via global settings Please use the Labbook icon in the MEA window for local settings and the icon in the Control View for global settings Please use the global settings which apply to all available MEAs or wells as well whereas changes in local settings will apply to a single MEA or a single well Changes in global settings will of course overwrite local settings If local and global settings
89. p is displayed 00 00 12 202200 In the screenshot above one well of a 256 9wellMEA with a 6 x 5 electrode grid is displayed Different layout grids are possible please see chapter Data Acquisition Settings Using a different type of a MEA for example a 256MEA with a 16 x 16 layout grid or a standard MEA with an 8 x 8 grid the layout will be adapted The Color Map Toolbar Click the Copy to Clipboard icon to copy the color map to the clipboard Click the Save Picture icon xd to save the color map as a picture in png format Click the Start icon start real time monitoring Click the Pause icon to stop real time monitoring Note Stopping the real time monitoring saves computer load 60 Cardio 2D Software 2200 us a 5342100 The electrodes on the MEA symbolized by circles Black circles indicate detection of a heartbeat on this electrode in the currently displayed cluster white circles indicate no detection of a heartbeat in the currently displayed cluster but the color map is completed by interpolation of data Click the Map Settings icon MEA 1 Map Settings The dialog Map Settings appears Select Map tab Data Map Color vl Invert Contour Black Aros 20 ms k Size Ww 260 x 290 Interpolation Bicubic Show Electrodes yl In this dialog you ca
90. parameter Please check the syntax EN 35 x The Detection Dead Time has to be longer than the Min Max Search Interval Please adjust the time 114 Theoretical Background I x valid license key Please contact licenseciemulkichannelsysbems cam Your license key is not valid Please contact Multi Channels Systems MCS GmbH Minimum Rise Time has to be shorter than Maximum Rise Time The minimum rise time has to be shorter than the maximum rise time 26 The Maximum Rise Time has to be langer than the Minimum Rise Time The maximum rise time has to be longer than the minimum rise time y Detection Dieadtime must be shorter than the shortest Post Heartbeat Interval of the amp verager The detection deadtime must be shorter than the shortest post heartbeat interval of the averager The Fost Heartbeat Interval must be greater than the Detection Deadtime of the OT Analysis The post heartbeat interval must be greater than the detection deadtime of the QT related Analysis 115 8 1 Cardio2D Manual Theoretical Background Cardiac Action Potential Over the last 30 years non invasive extracellular recording has developed into a widely used standard method The semi permeable lipid bilayer cell membrane separates different ion concentrations charges on the inner and outer side of the membrane Conventional methods me
91. patterns and controls that affect all three stimulus patterns First section The position indicator helps to navigate through the MEA electrodes 1 36 It shows the position of the mouse on the left MEA 1 or on the right MEA 2 The numbering of MEA electrodes in the 8x8 grid follows the standard numbering scheme for square grids The first digit is the column number and the second digit is the row number For example electrode 23 is positioned in the second column and the third row 39 Cardio2D Manual BEE BES BES Second section Click one of the three color coded Select Stimulus icons to select the respective stimulus pattern for further processing You can define the location of the selected stimulus by clicking on electrodes in the electrode panel or by drawing rectangles over the desired area Each stimulation electrode accepts one stimulus pattern at a time It is not possible to overlap patterns but to overwrite them with a new selection or to clear the sites To remove the currently selected stimulus from all sites please use the Clear selected stimulus sites icon created sites of the selected pattern will be removed To remove stimuli from single electrodes only cancel the current stimulus pattern selection first by clicking on the Select Stimulus icon again then click on the electrode you would like to remove Clicking the Define Stimulus icon will ope
92. pecu EMG ange 10 892 10 894 10 8 Gan Channel Overlay Resolution SampleRate 100006 gt i MEASettings 190 3 ade Main Window Overview The main window of Cardio2D is divided in two main windows On the left side is the Control Window and on the right side the MEA Window with the Field Potential ROI Selection Heartbeat HB Averager and the QT related Analysis tabbed page Control Window In the Recording control window on top of the left side the name s of the currently analyzed raw data and or result file is displayed You find additional information about the file in the table The Channel control window thereunder shows the MEA electrodes You can zoom to any electrode by clicking on the electrode The zoomed channel is displayed in the larger single channel view It is possible to define clusters in case of formation of a cluster Please see also the 2D tab in Tools Analyze dialog In the Settings file all information about the currently analyzed Cardio2D result or raw data file is displayed in a tree view 71 Cardio2D Manual MEA Window The window on the right side is divided into four separate window panes and has five tabs Field Potential ROI Selection Heartbeat HB Averager and QT related Analysis The setting of the tabs influences each of the separate windows in the tab Select Fi
93. potential signal Local means the first detected minimum or maximum global means the deepest minimum or highest maximum in the field potential within the Min Max search interval Please see the picture 2000 1000 FP uv 1000 Local Global 2000 minimum minimum LII EIL RNC CORN 1 88 1 89 Time s Whether you select the local or the global minimum or maximum depends on the aim of the user If you are interested in the maximal slope of the action potential you have to select the first local minimum or maximum If you are interested in absolute minimal or maximal values you have to select the global minimum or maximum Please select the time used for searching the minimum or maximum of the field potential Min Max Search from the drop down menu range from 1 ms up to 200 ms You can set a HB timestamp to each detected field potential either to the maximum or to the minimum or to the maximal slope of the field potential This timestamp is visualized in the plot with a black dot and stored in the raw data file Please read also chapter Detection Parameter Click HB Average tab in Analyzer Settings Analyzer Settings X MEA 1 Analyzer Settings X HB Detection Averager 2p HB Detection Averager 2p Fre HE Interval 50 ms E Past HB Interval ms Pre HE Interval Post HB Interval 33m It is p
94. ppendix Technical Support Test Model Probe Contact Information Index 71 71 84 86 87 95 97 105 108 111 114 114 114 116 116 119 119 119 120 121 1 1 1 2 Introduction About this Manual This manual comprises all important information about the first installation of the hardware and software and about the daily work with the instrument It is assumed that you have already a basic understanding of technical and software terms No special skills are required to read this manual If you are using the device for the first time please read the important safety advice before installing the hardware and software where you will find important information about the installation and first steps The printed manual and help are basically the same so it is up to you which one you will use The help offers you the advantage of scrolling through the text in a non linear fashion picking up all information you need especially if you use the index and the search function If you are going to read larger text passages however you may prefer the printed manual The device and the software are part of an ongoing developmental process Please understand that the provided documentation is not always up to date The latest information can be found in the help Check also the MCS web site www multichannelsystems com for downloading up to date manuals and help files Welcome to Cardio2D and Cardio2D The Cardio2D softwa
95. puter Please read the MEA2100 System manual for detailed information E Data Acquisition Settings X Device MEA2TOO 00017 A Y Sample Hate T Range 2500 my Resolution Channel Diver Version MEA 1 MEA 100 10 MEA 2 MEA 100 10 Close Data Acquisition with the USB ME Systems The external data acquisition board USB ME256 System is a stand alone solution for acquiring data from up to 256 channels from up to four MEAs It can replace the internal computer hardware MC_Card The analog input signals are acquired and digitized by the USB ME256 System and the digital electrode signals are transmitted to the connected computer via universal serial bus High Speed USB 2 0 Thus it is possible to use any computer for data processing Recorded data is graphed analyzed and reviewed with Cardio2D and Cardio2D program The USB ME256 System supports 256 channels with an amplification factor 1 that means no amplification data acquisition only You can connect for example up to four MEA1060 amplifiers via 68 pin MCS high grade cables 11 Cardio2D Manual The USB ME128 System and the USB ME64 System are in principle the same devices as the USB ME256 System but with 128 or 64 electrode channels respectively ES Data Acquisition Settings X Device LIS B MEZ5E Card 749 02 Sample Rate ALAS Hange 4096 mv Version MEA 1 MEA oxo 100 10 hi MEA 2 MEA 10
96. r MEA amplifiers which can be operated independent from each other That means you can select the settings for each MEA separately but the starting and stopping of the experimental protocol is synchronous for all connected MEAs The design of the MEA tab is different depending on the type of the connected MEA 23 Design of the MEA window when connecting three MEAs and using a MEA with a 6 x 10 layout grid in MEA 3 tab Design of the MEA window when using one EcoFlexMEA36 with a 6 x 6 layout grid Design of the MEA window when using a 256MEA with a 16 x16 layout grid Cardio2D Manual MEA 1 MEA 2 MEA 3 AAEE az ILL ram zi zi dii Ix x 4 1 TICOTT zl lk lai Ji jl Jl Jul Ji di DP AL PET a ty NE StL ELL et et let tet et def fol ey 1 et re et et ee 1 2 el et ee ae BRE RRR REE REE fe HHHH Tet et et et ie 1 H ELI dot EL tot Je HHHH L HH ATTAT naag qH 5 dsl ist LLELTE ELILI
97. rameters For example you can adapt the figure to a special layout wanted for a special publication Please see also chapter General Software Features 98 Theoretical Background Color Map 4700 ps Ops Click the Copy to Clipboard icon to copy color map to the clipboard Click the Save Picture icon to save the color map as a picture in png format 00 00 12 202200 da ms e u Note Stopping the real time monitoring saves computer performance The color map displays the spreading of the excitation of a heartbeat from the pacemaker to the outer regions of the cell culture or tissue If the pacemaker is not on the MEA the earliest point of detection is used instead of the pacemaker The isochrones are indicated by black lines The colors indicate the time distance between the first signal and the outer regions of excitations The action potential runs over the heart cell culture or tissue or hole heart from the pacemaker to the outer regions The MEA electrodes mirror this excitation The propagation time is the longest possible period for passing of the excitation from the first excited electrode to the last excited electrode of one cluster Define the propagation time in the 2D tab of the Analyze dialog Please see below If the propagation time is too short you miss data for the complete heartbeat If the propagation time is too long the next heartbeat starts and the results are incorre
98. re consists of two main parts The recording part is called Cardio2D program It is for controlling the amplifier the stimulator and for scheduling and running the experiments The offline analyzer part is called Cardio2D The Cardio2D analyzer program is intended for reviewing and graphical representation of recorded experiments and for preparing the graphical results as far as possible for a publication Raw data is recorded in 24 bit resolution when using the MEA2100 System The other data acquisition systems record raw data with 16 bit resolution Important Information and Instructions Important Safety Advice Warning Make sure to read the following advice prior to install or to use the device and the software If you do not fulfill all requirements stated below this may lead to malfunctions or breakage of connected hardware or even fatal injuries Warning Obey always the rules of local regulations and laws Only qualified personnel should be allowed to perform laboratory work Work according to good laboratory practice to obtain best results and to minimize risks The product has been built to the state of the art and in accordance with recognized safety engineering rules The device may only be used for its intended purpose be used when in a perfect condition Improper use could lead to serious even fatal injuries to the user or third parties and damage to the device itself or other material damage Warning The device a
99. re or tissue The isochrones are indicated by black lines The colors indicate the latency distribution over the MEA Red color represents the pacemaker when using the color map jet inverse The latency velocity window shows the latency of the heartbeat in units of us or the velocity in units of mm per ms Both parameters are measured between the first and the last detected electrode in the range The current absolute values are displayed in red letters above the plot 20 Cardio 2D Software Main Menu ES rardio2D Experiment Tools Settings Help File ES Cardin2D Experiment Tools 5 Open Template Save Template File Tools Help Open Raw Data Restore Defaults Open Result File Save Result File Save Defaults Exit Menu to open or save templates to restore and save defaults and to exit the program If you design experiments for a certain purpose and you want to reuse the experimental schedule please save the file as a template You can reuse the template file as often as necessary You can save as much templates as needed You can also save local and global settings as a template The inscriptions appear as soon as you open the template and there is no need to fill in again Factory set defaults are the setting parameters which are used when starting Cardio2D for the first time Please enter your own defaults for example in Data Acquisition Settings dialog after instal
100. rei 14 24 34 44 54 64 4 84 Downloag i AATA p E com Test Conn Connection OK Apply List Mode Blanking Stim B LE 53 58 When using more than one MEA amplifiers Port drop down menu for example 1 defines MEA amplifier 1 COM 2 belongs to amplifier 2 and so on Press Test Connection to be sure that amplifier and the computer are correct connected Select Stim A or Stim B in menu Stimulation Enable the check box Blanking Select the desired stimulation electrode on the electrode field The button will appear in blue when using Stim A or in red when using Stim B Click Download for downloading the information to the MEA amplifier 38 Cardio 2D Software 4 8 Stimulation with MEA2100 System Click Stimulate icon n When working with Cardio2D and any type of MEA2100 Systems with integrated stimulus generator STG the Stimulate dialog provides the possibility of stimulating via three different stimulation patterns on user defined electrodes Stimulate E AX MEA1 36 555 NIA gt b gt Close The dialog mirrors the layout of the currently used MEAs in this case a MEA2100 2x60 System with two slots for two 60 electrode MEAs Use the tool bar thereunder to define the stimulation sites Tool Bar The tool bar in the stimulate dialog is divided into three sections A position indicator controls for single stimulus
101. rlay plot The lower window Complete Stimulation Pattern shows the combined sequence of all stimulus pulses m 100 Shift 50 0 Periode 1000 a Phase 270 ES Cycles 7 151 us 1150 PP Amp m 23 ps 240 4 Marker 44 Cardio 2D Software Stimulus Pattern Rectangle Modulate the amplitude Amplitude mV of the ascending and the descend part of the pulse and the duration Duration ps of the rectangle pulse via up down boxes Setting the number of cycles to more than one enables the inter stimulus interval ISI us up down box Additionally the Arrow button will be enabled Amplitude mV 100 100 Duration ps 1000 a ES 1000 ES Cycles 1 at ISI ns E When using more than one cycle you can click onto the arrow symbol for additional options A highlighted parameter field appears to modulate the amplitude and the duration of the rectangle pulses in relationship to each other again independently for both parts of the pulse and for the duration between the ascending and descend arm The window Single Pattern shows the rectangle pulse in an overlay plot The lower window Complete Stimulation Pattern shows the combined sequence of all stimulus pulses Amplitude mV 50 160 Duration lt 2000 1400 1500 E Cycles 3 ISI us Amplitu
102. rovided primitive is a flat line stimulus To adjust the duration please use the respective up down boxes Hour minute second millisecond and microsecond Define the value of the pattern in the Amplitude mV up down box h m amp m Duration 0 0 1 0 Amplitude mV 0 43 Cardio2D Manual Stimulus Pattern Sine Wave Modulate the amplitude PP Amp mV the period Period us the shift Shift and the phase Phase of the sine wave pattern via up down boxes Setting the number of cycles to more than one enables the inter stimulus interval ISI us up down box Additionally the Arrow button will be enabled Amp mV 100 Shift Periade us 1000 ES Phase 1 ES Cycles 1 c 151 us When using more than one cycle you can click onto the arrow symbol for additional options A highlighted parameter field appears to modulate the amplitude and the period of the sine waves in relationship to each other In this example increases the amplitude of each sine wave at 23 mV in comparison to the sine wave before Choosing a negative value from the up down box the amplitude will decrease in comparison to the wave before Analogous you can adjust the value of the period in us in relation to the wave sequences The window Single Pattern shows the sine waves in an ove
103. ry of Multi Channel Systems MCS GmbH always apply The operator will receive these no later than on conclusion of the contract Multi Channel Systems MCS GmbH makes no guarantee as to the accuracy of any and all tests and data generated by the use of the device or the software It is up to the user to use good laboratory practice to establish the validity of his findings Guarantee and liability claims in the event of injury or material damage are excluded when they are the result of one of the following Improper use of the device Improper installation commissioning operation or maintenance of the device Operating the device when the safety and protective devices are defective and or inoperable Non observance of the instructions in the manual with regard to transport storage installation commissioning operation or maintenance of the device Unauthorized structural alterations to the device Unauthorized modifications to the system settings Inadequate monitoring of device components subject to wear Improperly executed and unauthorized repairs Unauthorized opening of the device or its components Catastrophic events due to the effect of foreign bodies or acts of God Operator s Obligations The operator is obliged to allow only persons to work on the device who are familiar with the safety at work and accident prevention regulations and have been instructed how to use the device are professionally qualified or have specia
104. se key To enter the following dialog select License Key in menu Help Please type the key into the window Please enter license key and confirm the key with Set License Please enter license key Successful registered Double click the Cardio2D exe on the installation volume The installation assistant will show up and guide you through the installation procedure Follow the instructions of the installation assistant Note The hardware driver is installed or updated automatically Installing the Cardio2D Analyzer Software Double click the Cardio2D exe on the installation volume The installation assistant will show up and guide you through the installation procedure Follow the instructions of the installation assistant You will again be asked for a path to put the log and data files Cardio2D does not produce raw data files result files only Raw data files created by Cardio2D are never changed by Cardio2D but analyzed and saved as result files It is advisable to use the same subfolders as used by the Cardio2D Software and Hardware Installation 3 2 First Use of Cardio2D after Installation Double click the Cardio2D program icon The Data Acquisition Settings dialog appears Driver Version Data Acquisition Settings Device 743 Sample Hate 20 kHz Hange 4096 mv ads Resolution Channel MEA 1 MEA 8 8 100 10 You have to select the dat
105. se of support 26 Cardio 2D Software 4 2 Control Window Control View Control Window Recording 2 Test E sperment stop Experiment start Recording 0 Apply Recording 1 Test pM Recording 2 Apply 100 pki Recording 3 Test 100 phl Recording 4 Apply Recording 5 Test 1 nM Recording Apply 10 Recording 7 Test 10 ntl Experiment stop Log File The Control Window is divided into two window panes The Control View controls the experimental protocol and the Log File documents the experimental proceedings 27 Cardio2D Manual 4 3 Control View In the Control window you can enter the settings for setting up and running an experiment If more than one MEA is connected each MEA is represented via a tabbed page in MEA Window In Control View it is possible to set different parameters for each MEA and to start and stop all MEAs simultaneously 28 4 4 4 5 Cardio 2D Software Viability m Stop Click Viability The Viability icon wv changes into Stop icon Sto and the duration of the viability test is counted up in hours minutes and seconds Stop the viability test manually by clicking on Stop icon a Stop Hi utm ig Running a viability test Before you start recording a viability test is recommended The data is not recorded The viability test monitors whether the MEA electrodes are acceptable for recording and the cell culture or tissue is fine Y
106. se select blanking feature in the MEA Select program please see chapter MEA Select Blanking Blanking Click the Start icon If your experimental schedule will always need the same pulse paradigm please store the settings in File menu Save Defaults This way the settings appear in a new file and you do not have to type in them again Stimulus Generator STG It is recommended to use a stimulus generator STG of the 2000 or 4000 series from Multi Channel Systems MCS GmbH Important When working for example with four amplifiers with blanking circuit BC you have to connect a STG4004 or a STG2004 The TTL blanking signal and the stimulation pulse must be synchronized that is you have to connect the stimulus pulse of channel 1 on MEA amplifier 1 to the TTL blanking pulse of Sync Out 1 of the STG channel 2 to Sync Out 2 channel 3 to Sync Out 3 and channel 4 to Sync Out 4 respectively When using an USB ME256 device for data acquisition you can connect up to four MEA amplifiers You define the stimulation settings for each of the connected MEA amplifier independent from each other and you can start and stop the stimulation independent from each other Amplifier without Blanking Circuit BC The stimulation electrode is manual selected directly on the amplifier via connector pins around the MEA You will see the answer of the stimulation during data recording because of the stimulus artefacts and the spreading of the fi
107. settings as default please enter File menu Save Defaults ES Cardio2D Experiment Tools 5 Open Template Save Template Restore Defaults Save Defaults Exit Data Acquisition Hardware Caution You have acquired a high performance data acquisition and analysis computer Do not modify the system do not install new hard or software or another operating system without asking MCS or your local retailer for advice Cardio2D do not install virus scanners or firewalls because these programs are known to interfere with the data transfer to the hard disk MCS cannot guarantee that a modified system is fully operational Even data loss may occur Warning The operating system settings of the data acquisition computer were preconfigured by MCS and should not be changed by the user Changing these settings can lead to program instabilities and data loss Please read chapter Data Acquisition Settings in First Use of Cardio2D after Installation 10 Software and Hardware Installation Data Acquisition with the MEA2100 Systems The MEA2100 recording system is an all in one solution consisting of headstage and interface board The MEA2100 System with integrated amplification data acquisition online signal processing and integrated stimulus generator You can connect one or two headstages to the interface board The MEA2100 System uses an USB 2 0 connector per headstage to transfer digitized data to any com
108. ssue The isochrones are indicated by black lines The colors indicates the time distance between the first signal and outer regions of excitation Red color represents the pacemaker when using the color map jet inverse The color map corresponds with the single channel view and with the MEA electrode field 1000 us 8967400 Click the Copy to Clipboard icon to copy the color map the clipboard Click the Save Picture icon Izd to save the color map as a picture in png format Note Stopping the real time monitoring saves computer performance 91 Cardio2D Manual Click the Map Settings icon The dialog Map Settings appears Click Map tab MEA 1 Map Settings Data Color Jet Invert Contour Black ATONE Dm Size a H 290 290 Interpolation Bicubic Show Electrodes y In this dialog you have to define the settings for the color map Color Map The usual used color map for cardiac issues is the Jet Invers map but you can also choose Hot Cool or Grey maps To remove the colors from the map click Off The isochrones will remain only Contour The drop down menu is for changing the color of the contour for the isochrones into Black Jet Hot Cool and Grey Or you can remove the lines with Off Adjust the number of the contour lines from the numeric box beside up to 100 contour lines are possible Arro
109. t the experiment Important If you remove electrodes from the screen data streams of these electrodes are not saved in the data file 40 Cardio 2D Software Stimulation Pattern To open the Define Stimulus 1 dialog please select the first Select Stimulus 1 icon The Define Stimulus icon in the Stimulation window will be available Each stimulus can be defined independently so the dialog corresponds with the previous selected stimulus pattern 1 2 or 3 The dialog for the settings of the stimulus patterns are build analog here for example the Define Stimulus 1 dialog Define Stimulus 1 Amplitude m 100 100 Duration 300 E 120 450 E Cycles at ISI us E Parameter Settings Single Pattern Primitves J Marker Complete Stimulus Pattern Amplitude 0 2 Offsetimv 0 2 oo E LL of 3 The Define Stimulus dialog is divided in an upper and a lower section The upper part includes the icons for the provided stimulus patterns such as flat line rectangle pulse ramp or sine waves The last icon represents biological signals for example which can be imported into the Cardio2D software The lower section of the dialog shows the Complete Stimulus Pattern and a tool bar The settings in the tool bar influence the complete sequence of the stimulus patterns which are immediately displayed in the window for the Complete Stimulus Pattern
110. ted raw data file The numbers of the electrodes refer to their position in the grid to rows and columns Using a different type of a MEA for example the layout will be adapted If you have different clusters of cells with different pacemakers on the MEA you have to analyze each cluster separate Mark the electrodes belonging to the first cluster by using the Add Channel icon Lj to select channels Deselect the other electrodes by clicking the Remove Channels icon Run the analysis and save the results After doing this you can mark the next cluster for running a second analysis Settings File MEASsttings a MEAID Selected Layout RegularGrid 3ridH eight Grids pace arid5 pace GridColurnins ows Channels E Analyze H MapFieldPotential MapLatency Datalnterpalatian Lhannellnterpaelatian Min Min amp uta Mapinterpalation In the Settings file all settings of the currently analyzed Cardio2D result or raw data file are displayed in a tree view The window contains all information saved during recording the file and you can read any information you need for analyzing the file It is not useful and therefore not possible to change these information in a Cardio2D raw data file If you re analyze a raw data file with new parameters you can save the new results in a new Cardio2D result file MEA Window Cardio2D Th
111. terns click on the icon Create Movie A second red bar appears Now the function of the red bars is similar to the function of the red bars in single channel view The two red bars here limit the time span for the movie a region of interest too Click on the point in the middle of the bars The mouse pointer turns in an arrow with two arrow heads and you can vary the time span between the two bars Click on any other point of the red bars The mouse pointer turns in a cross with four arrow heads and you can move both bars to the section of the field potential you want to save in a movie Click the icon Create Movie Click on the icon Start gt 40 E 34 20 0 20 Th N P RON T 40 IN I amp A 22 85 Time 500 py TERT 3 1000 us m While recording the movie the blue arrow indicates the progress of recording You can disrupt the movie by clicking the icon Pause Movie The inter picture timing 1300 us iq us allows adjusting speed and duration of a movie generated by using the above described functions 90 Theoretical Background Color Map The color map displays the field potentials on the MEA electrodes at a time stamp defined by the channel overlay display It shows the excitation of a heartbeat from the pacemaker or the earliest time point detected if the pacemaker is not on the MEA to the outer regions of the cardiac cell culture or ti
112. ting the online help for Cardio2D and for displaying the dialog About Cardio2D ga about CardiozD Cardio2D Version 1 5 3 Multi Channel Systems GmbH MES Copyright MCS 2008 2012 Close The dialog shows basic information about Cardio2D Note Please keep in mind that the information of this dialog is necessary in case of support 83 Cardio2D Manual Control Window Cardio2D The control window of Cardio2D is divided in three separate windows The Recording control on top the Selection control view and the Settings file below Recording Control C Emm 1 wed C Birgit Cardio2D 5 imD ata amp stemieoleMEAR 10 Type Dose Compo Duration Channel Control Control Window le Bus x Ere Bar 28 38 45 58 Mud E Cardio2D Results Labbook Environment Experiment Stimulate DataSource MEAS ettings Analyze H HBDetectMode HBTimeStampT ppe ime Settings File L etectS actor DetectD eadT ime DetectSearchinteryal PrapagatianP ath Prapagationll rit Propagation T imeh PropagatianPathElectrodelD s gt MapFieldPatential MapLatencu MovieFieldPatential Maviel atenc 84 Theoretical Background Recording Control of Cardio2D L sBirgirCardiogD Daten A0S MES
113. trode view displays electrode activity of all electrodes in the layout The Single Channel View displays an user defined single channel to have a closer view You can display the FP field potential the HB heartbeat and the Ave average The Color Map displays the propagation of a heartbeat from the pacemaker to the outer regions of the cardiac cell culture or tissue The Latency Velocity view displays the latency or the velocity between the first and the last detected electrode channel 54 Cardio 2D Software The MEA Window changes depending on the connected hardware If you connect for example an USB MEA256 System for data acquisition you can use it together with a 256 9wellMEA and each of the nine wells is displayed and controlled separate in nine MEA window tabs MEA A to MEA 1 MEAC MEAD MEAE MEAF 5 MEAH MEA If you connect for example an USB ME256 System for data acquisition it is possible to connect up to four MEAs MEA 1 to MEA 4 MEA 1 MEA 2 MEA 3 MEA 4 You can switch between the different window tabs to set different setting parameters in the four views of the MEA Window independent from each other Via Control Window you start and stop the experimental protocol of the connected MEAs simultaneously 4 14 MEA Electrodes The MEA electrodes window allows real time monitoring of field potential data The electrode channels are arr
114. two points of a line The bilinear interpolation uses four points of the nearest vicinity to calculate the interpolated point The bicubic method uses eight points to calculate the interpolated point The interpolation method of the nearest neighbor uses the mean value of four or five points to calculate the interpolated point Show Electrodes Click the check box for displaying the black circles indicating the electrodes in the color map 62 Cardio 2D Software Select Data tab in Map Settings dialog MEA 1 Settings ata Map Interpalatian Missing Data MearestMeighbor Missing Channel MearestMeighbar Show Time Stamp hhmmss ms hhmmss Interpolation In the Map tab of the Map Settings dialog the interpolation parameter refers to points between measured data In the Data tab the interpolation parameter refers to the data generating the map Default is the interpolation method Nearest Neighbor Please see above It is possible to interpolate Missing Data or Missing Channel Show Timestamp You can add the heartbeat timestamp to the color map using the option Show Timestamp Different settings are offered for the actual timestamp which is displayed below the color map For displaying heartbeats for example a timestamp in milliseconds ms will be enough for potential spreading you can use the timestamp option hours minutes seconds and microseconds hhmmss ys Click the Record Movie i
115. ulation in ms in the second up down box to define the time interval in which the slope of one point of the signal is calculated Please read chapter Automated Waveform Detection in Analyzer Setting This feature works like a filter because the remaining noise is reduced QT related Interval 327 ms M MM 0 210 400 Bt The left window shows the different recording phases of the selected electrode starting with the smallest amount of compound which is the reference by default 0 uM The molarity of the compound and the number of detected and averaged heartbeats are displayed in the green rectangle in the upper left edge The blue bar indicates the end of the QT like Interval calculated by the computer If you change the level manually the bar turns into red color The duration time of the QT like Interval in milliseconds is written beside the bar EERE 0 200 400 Bt Click onto the green or red rectangle to select a recording for the calculation of the dose response curve 109 Cardio2D Manual Dose Response Curve The right side shows a diagram of the dose response curve if the recorded data allow to calculate it m ze I C 100 1000 10000 100000 Dose pM Absolute The curve is available in Normalized or in Absolute form the x axis is logarithmic In Normalized plots t
116. ve result files to open raw data files and to exit the program Cardio2D generates different types of files the Raw Data file c2d the Result file c2r and the Settings file c2s Raw data file and the associated settings file always belong together in the result file the settings file is included The log file is not automatically generated but by users command If the Averager tool is activated during recording you generate additionally an Averager file c2a The Cardio2D Analyzer program deals with raw data and result files It is not possible to change the data of a raw data file which was recorded with Cardio2D But you can re analyze a raw data file with Cardio2D Analyzer as often as necessary and save the new information in a result file When a result file is re analyzed no data will be overwritten but a backup file of the old result data is generated backup Additionally an analysis file can be generated when a result file is analyzed with the heartbeat average feature c2a Tools ES rardio2D File Tools Help Analvze Raw Data Menu to open the Analyze dialog Click Analyze Raw Data 77 Cardio2D Manual Click HB Detection tab in the Analyze dialog da analyze X HB Detection HB Average 2D Minimum Rize Time 100 pus Maximum Rise Time 1 ms Fixed Amplitude Signal Dependent 10 StdDev Detection Deadtime 5U ms Min Max Global Min
117. ws The feature is not yet implemented in this version of Cardio2D As soon as it will be implemented you find a Read Me on the MCS web site where you will be able to download the new version of Cardio2D Minimum Maximum You have to define the minimum and the maximum of the scale of the color map That means you define the time frame in which the excitation pattern are displayed in the color map Usually you start with zero in the minimum that is the pacemaker The excitation pattern will run from the pacemaker over the map to the maximum The widest selectable time frame for the color map spans from 0 us in the minimum to 10000 ms in the maximum To change the colors of the map please use the first drop down menu Color Map 92 Theoretical Background Auto Click the check box Auto in Maximum option The Minimum is zero you are not allowed to modify it in Cardio2D The Cardio2D program automatically finds for the maximum of each heartbeat and adapts the scaling of the color map It is recommended to use the auto scaling function Size W x H You can modify the size of the color map Interpolation Interpolation is a mathematic method to calculate missing information between two points of a line The bilinear interpolation uses four points of the nearest vicinity to calculate the interpolated point The bicubic method uses eight points to calculate the interpolated point The interpolation method of the nearest neighbor uses

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