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User Manual - Thermo Fisher Scientific

1. cceeseseseseseseseseseeeeeeeseceseseseseneaeeeeeeeeeteeeeseseaeaeneeeeeees 49 EVOS Onstage Incubator Control Unit Instrument Panel ccssessssssesesesescseseneseneneneaeaeaeaeaeaeeeseeeens 49 EVOS Onstage Incubator Stagetop Environmental Chamber 50 Installing the BVOS Onstace Incubator eause tears actoecatina ua 51 Using the EVOS Onstage meubatome ee made benne 57 General Careof the EVOS Onstace OUAIS Re a dde 59 Steril zat On Procedures nus ed As te te a a barn cereale 59 PO DCMOIX CS al ely APE EE EE EEES 60 Safety Conventions Used in this Document sessirnir aii imitant 60 SYMPOIS On LTS TUMMEN iara se elects lost vc Peace cae nse vn oe nnn evs nd an inst 61 Dalety Labels On MS CN CIS SSSR nn indienne naine 63 General IMS EMEA SR dt n TERE a NA suing uanstougad TRS 64 Safety Requirements for EVOS Onstage Incubator ss 65 PETS TY CAA SG Re ee es peste oats oat aie apes ye mm pe omen ne Or nee 66 Chemical Waste SAR Re dE ie entente 67 PIC STD eee ne een te eee ne ee re eo et Cn er nes 68 Physicak Hazard SAL E ne A Ann Sedat aga ae ones Rene qe 69 Biological Hazard SA Est ee T E eaeoetoemtomaarentels 69 Safety and Electromagnetic Compatibility EMC Standards ss 70 Documentation and Support cceeccecceeeeeeeeeeeeeeaeeeeeseesaeeseesaesaeesaesaeeeeesaneneesensaneneesaes 71 OPA iro b 1 0 0 0 E SR a a A Ce A de te 7A About This Guide Audience User Attention Words Safety Alert
2. X Note Store the Shipping Restraint in your accessories box for future use Turn ON the 1 Turn the instrument power switch located on the right side of the instrument EVOS FL Auto base see page 45 to the ON position Imaging System The automatic X Y stage will begin to move within a few seconds of the instrument being powered on Turn the computer and monitor ON When the computer shows the Windows desktop and the X Y stage of the instrument has stopped moving double click the EVOS logo next to the start button on the desktop to start the EVOS FL Auto software 4 The EVOS loading screen will be displayed while the software is starting up This may take a minute 5 Once the Home page see page 7 is displayed the EVOS FL Auto Imaging System is ready to use Home Page Home page is displayed when EVOS FL Auto software is launched It allows you to choose between the options listed below QUICK IMAGE CAPTURE Help me capture an image quickly Capture a single image while controlling the scope directly SCAN TIME LAPSE Scan an area and capture multiple images Build your own time lapse movie to stick together e Quick Image Provides step by step instructions for capturing an image in the fastest way possible See page 8 for more information e Capture Captures a single image while allowing the direct control of the instrument See page 10 for more information e Scan Scans an area to acquire m
3. BMP TIFF and PNG Save As Allows you to choose between Color or Monochrome Include Image Annotations OL 9 SAVE IMAGE 2 Include Image Annotations Q Allows you to include image annotations in your saved image Save Underlying Channels Allows you to include the underlying channels in your saved image Save Image Saves your image using the selected options amp Note The Default option saves images in 8 bit color in TIFF format with the image annotations included To save a 16 bit image select TIFF and ensure that the Color option is deselected File types JPEG and PNG as well as images of all types with the Color options engaged only save at 8 bit depth 3 Editing and Analyzing Images Overlay Overlay Tab Controls The Image gt Edit gt Overlay tab allows further adjustments to the brightness and contrast in each channel before the captured image is saved Each channel can be turned ON and OFF independently and the brightness and contrast for each channel can be adjusted separately using controls described below ON OFF buttons Switch the light IMAGE SCAN TIMELAPSE SYSTEM cubes corresponding to the fluorescent capture el save review sertincs channels and the transmitted light from SC UT mesure count aurocounr the condenser ON or OFF Channel buttons Select the channel for adjusting the brightness and contrast The channel being adjusted is
4. Cover the parfocality grid slide with the light box supplied with the EVOS FL Auto Imaging System but remove the light box cover 5 Click Next to proceed with finding the parfocality offsets for the combination of the objective and camera selected 6 On Find and Focus use the controls to bring the grid pattern on the parfocality grid slide into focus and then click Next again Repeat the procedure for desired objective and camera combinations After completing the calibration procedure click Finish 37 Calibrate Vessel Alignment 38 _ Go to System gt Basic tab page 30 and then click Calibrate Vessel Alignment to open the Vessel Alignment Tool which provides step by step instructions for adjusting the alignment positions of the current vessel Adjust the lighting and focus on the vessel using the Light slider and Focus controls Press the TOP button to move to the current top position Adjust the view so that the top edge of the vessel exactly touches the horizontal crosshair on the screen Click Save to lock in the new top position Repeat the procedure for each Upper Left Well DIAMETER Lower Right Well AUTO FINE SNAP TO LAST AND SAMPLE COARSE eme QUICK remaining parameter LEFT RIGHT and BOTTOM Once you have calibrated the vessel alignment for each parameter click Finish Changing the EVOS LED Light Cube Changing the LED Light Cube
5. Create Folder button Creates a new folder in the current location Selected File Displays the name and date of creation for the selected file Rename button Allows you to References rename the selected file Delete button Deletes the selected file Save Image Saves the image in the desired location For more information see page 13 Timelapse_OGrady 20130418 Mous eBrain_ACD_Posit 20 4 Advanced Operations Image Settings The Image gt Settings tab allows you to customize the user interface of the EVOS FL Auto Imaging System for your needs by selecting up to four Hot buttons for the Image gt Capture tab and by assigning the pseudo colors for each channel when the Monochrome camera is used Image Capture Hot Buttons are user programmable WAGE SCAN TIMELAPSE SYSTEM Programmable image capture Se that serena a CT com saw CTT D imaging system You can select up to four IP he Hot Buttons to be displayed on the Eu Image gt Capture tab image Capture Programmable Buttons Button 2 Capture All Channels X Button 3 e Capture All Captures an image using ee each of the installed light cubes e Capture Captures the current image on the screen e Preset Utilizes the capture options Corie CO Giang selected with the Quick Image gt Edit Launch Wizard see page 8 e Transfection Creates a two image overlay using one fluorescent image Edit a
6. click Delete All Annotations Count Count Tool Count Tool Controls The Count tool located in the Image gt Edit gt Count tab allows you to manually mark items onscreen using up to six separate labels As you tag each item to assign one of the six labels thus grouping similar items together the system keeps a running tally of the counts with percentages for each label assigned amp Note You can use the Count tool on both newly captured and saved images and document the results of your count by saving the tagged image with the Count tool displaying the totals IMAGE SCAN TIME LAPSE SYSTEM _ Tags Add Activates the Add mode to tag items onscreen using the selected Label see below Select Activates the Select mode to select existing tags onscreen Zoom Activates the digital zoom sense function for a closer look at live or captured images Labels Select the label used for tagging on screen items Each label row features a text field for entering a label 0 0 name and displays arunning tally of the count with percentage Clicking in the trash bin icons deletes the tags on the screen for that label set 0 0 9 o 0 0 10 0 0 0 Total Displays the total count of all labels assigned Delete All Tags Misc V2 Show Tags Delete All Tags Deletes all existing gt js ETES tags s Save Image As Screenshot SAVE IMAGE 3 Show Tags D
7. indicated by the blue color of the corresponding button In the example on the right the RFP channel is selected for adjustment Brightness and Contrast sliders Adjust the brightness and contrast of the image in the selected channel Reset All Channels Returns the brighness and contrats settings to their rere previous value Clear Channel Clears the memory buffer for the selected channel RESET ALL CHANNELS CLEAR CHANNEL Save Image Saves the image in the desired location For more information see page 13 SAVE IMAGE 6 amp Note Captured images can be saved in JPEG BMP TIFF and PNG formats For more information and additional save options see page 13 Measure Measure Tab Controls The Image gt Edit gt Measure tab is used for adding annotations creating histograms for visualizing data and drawing regions of interest ROIs in the captured image prior to saving Annotations Wace scan TIMELAPSE SYSTEM Add Activates the Add mode to add al seven semnos an annotation using the Annotation tools Ome eee NEC auro rO see below ara To add an annotation click Add select an Annotation tool and then click the area of the image where you want to add the annotation Select O Activates the Select mode to edit the properties of an existing eo annotation or the contents of a text box Zoom Activates the digital zoom function onscreen allowing a clo
8. 40 90 F e Relative humidity range 30 90 Q IMPORTANT Do not position the instrument so that it is difficult to turn off the main power switch located on the right side of the instrument base see page 45 In case of an instrument malfunction turn the main power switch to the OFF position and disconnect the instrument from the wall outlet Set Up Unpack the Monitor Unpack the Computer Unpack the Instrument og Se tS So ee Se IY e Open the case and remove the monitor and accessories If a VGA cable is attached to the monitor take it off Remove protective covering from monitor Plug power cord into monitor Plug USB cable into monitor Open the box and remove the keyboard Unpack the keyboard from its box Unpack the computer Unpack the mouse and power cord from the accessory holder Plug in mouse keyboard and power cord Plug the USB cord already connected to the monitor into the computer Open the box and remove the accessory box Carefully lift the instrument out of the box holding it by one of the four handholds in the base see page 45 D IMPORTANT Do not lift the EVOS FL Auto Imaging System by stage or condenser arm Lift the instrument by using the handholds in the base Place the instrument on a flat level surface that will be free from vibration and leave enough room around it for the stage to move freely Remove the following from the accessory box located in the i
9. Auto Imaging System from our website The updated user guide covers the new software functionality when features are added Alternatively you can get the latest software and documentation updates from your local EVOS distributor or by contacting Technical Support page 71 Plug USB flash drive into computer RER E Click System tab in EVOS FL Auto site software and then select the CONTACT LIFE TECHNOLOGIES FOR SERVICE Service tab Serial Number ce jii Click Update Software in the service tab A verification progress bar should appear amp Note If a missing update notification pops up be sure the USB flash drive with the update file is fully plugged in Click OK and then click the update button again After file verification an update UPDATE permission dialog will pop up Please confirm that you wish to re install software revision 22029 Check the revision details and click Yes to start the update Yes No A Windows Setup dialog box will pop up Click Next to proceed with the software update The screen will display the update progress When the update is complete the Windows desktop will appear Click the EVOS FL Auto icon in Windows to start the software IMPORTANT Do not power off unplug the USB flash drive or add or remove any devices during the update 39 Calibrating the EVOS FL Auto Imaging System Calibrate the Field of View 36 dk 10 11 Go
10. Incubator see page 58 Configure Gas Connections is used for configuring the gas connection ports on the control unit of the optional EVOS Onstage Incubator see page 57 amp Note The Calibrate Oxygen Sensor and Configure Gas Connections buttons are only available when the optional EVOS Onstage Incubator is installed with the EVOS FL Auto Imaging System See page 47 for more information on the optional EVOS Onstage Incubator Capture Focus To use this feature and recalibrate the Z plane for your vessel follow the steps Nominal below 1 Navigate to Image Capture tab adjust the focusing sliders to get the sample in focus and capture the image 2 Go to the System gt Basic tab and click Capture Focus Nominal 30 Network Tab Helpful Hints Using the System gt Network tab you can log the EVOS FL Auto Imaging System onto a Windows SMB network via an Ethernet cable connection and save captured images directly to shared folders on the network amp Note SMB is the only supported protocol No other protocols such as HTTPS FTP or WebDAV are currently supported If you are connecting to a Linux server it will have to use Samba for EVOS FL Auto Imaging System to find it Contact your network administrator for help if a physically connected EVOS FL Auto Imaging System cannot find the SMB network The EVOS FL Auto Imaging System IMAGE SCAN TIMELAPSE I SYSTEM recognizes the netw
11. Onstage Incubator Gas Line Cat no AMEP4732 e Push to connect gas line adaptor 3 each e Standard head open end wrench e Hex screw driver e Power Cord Type A North America Note Note that a country specific power cord must be ordered separately in regions not using the Type A power plug 47 Technical Specifications Physical Characteristics Hardware 48 Note Specifications of the Onstage Incubator are subject to change without notice Refer to the EVOS FL Auto Imaging System product page at www lifetechnologies com evosflauto for the latest product information Stagetop Environmental Chamber Control Unit Height 25 cm 9 7 in 37 cm 15 in Depth 19 cm 7 6 in 16 cm 6 3 in Width 3 7 cm 1 5 in 20 cm 7 9 in Weight 1 5 kg 3 3 1b 10 kg 22 1b Temperature range Ambient to 40 C 0 1 C Humidity gt 80 relative humidity RH at 37 40 C CO range 0 to 20 O range 0 to ambient Operating power 100 240 VAC 1 8 A Frequency 50 60 Hz Electrical input 24 VDC 5 A Compatible vessels Multi well plates 35 mm Petri dishes T 25 flasks Gas input ports Air or air CO premix CO only and N only max 50 psi input Stagetop environmental chamber accessories Master Stage Plate Vessel holder for multi well plates Lid retainer shield EVOS Onstage Incubator Exterior Components Control unit Onstage incubator heated hose a Aii T VAA Wi Hose heater connector
12. TIFF Folder NETWORK3 EVOS Timelapse_OGrady P Set beacons 4 Set beacons on locations in your Time Lapse Time Lapse Time Lapse sample vessel to be captured during 20130404_Chuc 20130404_Chuc 20130404 Chuc k_60X 2013 04 k_40X 2013 04 k_60X 2013 04 the time lapse session a Click Power to ON position b Select Pan mode and navigate rics tae aa aa dine aces to first Region of Interest ROI KGOX 201304 k 60X 201304 k 20X H2BRFP c Set focus and lighting which will be used on all subsequent beacons Time Lapse Time Lapse Time Lapse 20130405 Chuc 20130405__Lyso 20130407_Tran k_40X 2013 04 2013 04 05 13 s_20X 2013 04 Note Selecting autofocus when setting up the time lapse routine will cause autofocus at each beacon d Select Add Beacon mode and then click on ROI on the live image e Repeat steps b d until you have set up all desired beacons Note To review all set beacons use the Select Previous or Select Next beacon buttons 28 Reviewing Time Lapse Create Scenes 5 Using the Create Scenes function you can change the conditions during the period of the time lapse Select the location to save the routine Select whether to automatically create a video and place Life Technologies watermark on video 8 Run the Time Lapse routine The Time Lapse gt Review tab utilizes the same folder system as the Image gt Review tab page 20 for easi
13. To customize your EVOS FL Auto Imaging System you can add and remove LED light cubes to fit the instruments functionality to your own specific research needs For instructions on how to change the Light Cubes see below For a complete list of available light cubes and to inquire about custom light cubes go to www lifetechnologies com evos or contact Technical Support see page 71 SCAN TIME LAPSE SYSTEM 1 On the Image Capture tab select the channel of the filter cube you want to remove replace 2 On the System Service tab click Move XY to Filter Cube Access Position PA ee a A dA 20000000 90000000 VESSEL EXPERT 00 0 00 900000 eee oe If no cube is installed proceed to Step 7 eee 000 all e Use the light cube tool to loosen the two slotted screws as shown below F IMAGE SCAN TIME LAPSE SYSTEM m basic uerwonx JS 2 5 Screw the threaded end of the light cube STE ES tool into the hole in the center of the light This EVOS Auto cube as shown ion 22 Objective Lens AMG 4X PlanPH AMEP 4632 X AMG 20X Pian Fi AMEP 462 20X Plan Fi AMEP 4624 x 6 Use the tool to tilt the light cube slightly toward you and lift out gently and then remove tool from cube 7 Attach the tool to the new light cube and A lower the cube into position so that the TO electronic connection aligns properly facing the back of the microscope and the
14. Words This user guide is for laboratory staff operating maintaining and analyzing data using the EVOS FL Auto Imaging System Two user attention words appear in Life Technologies user documentation Each word implies a particular level of observation or action as described below Note Provides information that may be of interest or help but is not critical to the use of the product Q IMPORTANT Provides information that is necessary for proper instrument operation accurate installation or safe use of a chemical Four safety alert words appear in Life Technologies user documentation at points in the document where you need to be aware of relevant hazards Each alert word IMPORTANT CAUTION WARNING DANGER implies a particular level of observation or action as defined below Q IMPORTANT Provides information that is necessary for proper instrument operation accurate installation or safe use of a chemical CAUTION Indicates a potentially hazardous situation that if not avoided may result in minor or moderate injury It may also be used to alert against unsafe practices WARNING Indicates a potentially hazardous situation that if not avoided could result in death or serious injury T DANGER Indicates an imminently hazardous situation that if not avoided will result in death or serious injury This signal word is to be limited to the most extreme situations Except f
15. as desired during a count the EVOS FL Auto software will tag each item for the label selected To use digital zoom while counting click Zoom to select the Zoom mode and then double click the area onscreen you wish to enlarge To resume counting click Add to reselect the Add mode To move a tag click Select to activate the Select mode and then click and drag the tag to the desired location Left click anywhere on the screen to deselect it To delete a tag right click it You may also click Delete All Tags to delete all tags for all labels To save the image with the tags showing the labels counts and percentages as displayed in the Count tool select Save Image as Screenshot and then click Save Image to save the image in the desired location LS Note Deselecting the Save Image as Screenshot option will produce an image saved with the tags only Auto Count Auto Count Tool Auto Count Controls The Auto Count tool located in the Image gt Edit Auto Count tab is used for automatic counting of regions of interets ROI on an image Based on a histogram of the captured image and gated by signal intensity the tool uses an algorithm to mark and count the regions of interets ROJ on the screen see the example below Count button starts the Auto Count ann sitet mai procedure for the captured or live image a ME ET on the screen mine Count From File button opens a Count p
16. be automatically directed to the Image gt Capture tab see next page 4 Inthe Which Lighting field select the channel for previewing the sample and capturing the image You can only select a single channel for the preview but you may choose multiple channels for the capture overlay 5 Inthe What Magnification field select the objective to preview the image and then select the same or different objective to capture the image s Q IMPORTANT For best results preview your sample in a lower or the same magnification with which you would like to capture your image 6 Click Launch Wizard The system will move to the Image gt Capture tab which will display the preview choices selected in Step 3 LIGHT AND CAMERA POWER 5 u DAPI gt GFP TiRed TRANS No Image No Image No Image FIND SAMPLE AUTO FINE ON MO oy CAPTURE ALL cr COARSE 7 Adjust the illumination and the focus using the Light slider and the Focus controls You can make adjustments to each individual channel by first selecting it with the Channel buttons 8 Press Preset to utilize the capture options selected in Step 3 The system will automatically move to the Image gt Edit gt Overlay tab to allow for further adjustments to brightness and contrast using the Brightness and Contrast sliders D ON OFF ON OFF ON OFF DAPI GFP RFP CONTRAST 61 RESET ALL CHANNELS CLEA
17. chemicals in the instrument including liquid in the lines are potentially hazardous Always determine what chemicals have been used in the instrument before changing reagents or instrument components Wear appropriate eyewear protective clothing and gloves when working on the instrument VAR WARNING CHEMICAL STORAGE HAZARD Never collect or store waste in a glass container because of the risk of breaking or shattering Reagent and waste bottles can crack and leak Each waste bottle should be secured in a low density polyethylene safety container with the cover fastened and the handles locked in the upright position Wear appropriate eyewear clothing and gloves when handling reagent and waste bottles General Safety To minimize the hazards of chemicals Guidelines e Read and understand the Safety Data Sheets SDSs provided by the chemical manufacturer before you store handle or work with any chemicals or hazardous materials See Safety Data Sheets SDS e Minimize contact with chemicals Wear appropriate personal protective equipment when handling chemicals for example safety glasses gloves or protective clothing For additional safety guidelines consult the SDS e Minimize the inhalation of chemicals Do not leave chemical containers open Use only with adequate ventilation for example fume hood For additional safety guidelines consult the SDS e Check regularly for chemical leaks or spills If a leak or
18. cube sits squarely in place 8 Use the light cube tool to gently tighten the two slotted screws so that the screw heads sit flush with the ridges on the light cube IMPORTANT If the screws are not flush with the top of the light cube they may catch on the stage while moving and damage the system 37 Installing the Shipping Restraint Install Procedure 40 Use the Shipping Restraint whenever you package the unit for shipment If the unit is being hand carried and is not at risk of drops or excessive vibration there is no need to install the restraint If you need to remove light cubes or objectives remember to do so before installation of the Shipping Restraint 1 Start with the microscope turned on and the software running Navigate to the Service tab under Settings and click Move to Shipping Position A dialog box will appear detailing Step 1 Follow the instructions and click Move Wait while the microscope moves into position the mouse cursor will be busy during this operation Once the stage comes to a halt Step 2 will appear Align the pins with the holes on the mid plate of the stage and secure thumbscrew 1 Press Move to bring the top plate into position Step 3 will appear Secure thumbscrew 2 and press Move to bring the bottom plate into position Step 4 will appear Secure thumbscrew 3 and press Finish to advance to Step 5 Verify that all thumbscrews are secure Click Done and power down the in
19. each port 4 Turn on the regulators on the gas tanks The meters on the regulators show the tank fill on the right and gas flow on the left 5 Set the flow on the regulators as follows Do not exceed 50 psi of pressure e CO 5 psi e Air 40 psi e Nitrogen 40 psi 57 Calibrate Oxygen Sensor 08 Calibrating the oxygen sensor ensures that the atmosphere in the Stagetop Environmental Chamber is replenished with the appropriate gasses in the correct proportion 1 Goto System gt Basic tab page 30 and then click Calibrate Oxygen Sensor Oxygen Sensor Calibration This process will take approximately four minutes Please verify gas configuration before proceeding Which pure gas source would you like to use for the calibration process A co2 Begin Calibration Verify that your gas connections have been correctly configured Select the Pure Gas Source for your calibration Nitrogen or CO Click Begin Calibration The EVOS FL Auto Imaging System automatically calibrates the oxygen sensor for the proper functioning of the EVOS Onstage Incubator The entire calibration process takes approximately four minutes General Care of the EVOS Onstage Incubator When cleaning the EVOS Onstage Incubator use only optical grade materials to avoid scratching soft lens coatings Use the appropriate cleaning solutions for each component as indicated in the Sterilization Procedures below If liquid spills on the ins
20. the USB flash drive into the computer and select this button Error log will be automatically downloaded to be Calibrate Field of View sent to Life Technologies ee HS Microscope Use the drop down menus to select the correct objective for FL Hot Pixel Correction each objective turret position Correct Calibrate Parcentration Select to calibrate the objectives for parcentricity using a calibration slide Correct parcentration ensures that an object in the center of the field of view will stay in the center of the field no matter which objective is being used Move XY to Filter Cube Access Position Move to Shipping Position Calibrate Field of View Select to calibrate the field of view using a calibration slide see page 36 Calibrate Parfocality Select to calibrate the objectives for parfocality using a calibration slide see page 37 Calibrating the parfocality ensures that the sample stays in focus when the objective is changed FL Hot Pixel Correction Select to correct hot pixels on camera Requires covering stage with light box with cover to prevent stray light from entering optical path Date and Time Click to set the date and time Move XY to Filter Cube Access Position Clicking this button will move the stage to a position that will allow for easy installation of light cubes see page 39 Move to Shipping Position Follow prompts to properly install the shipping restrain for sa
21. to System gt Service tab page 32 and then click Calibrate Field of View to open the Calibration Wizard which provides step by step instructions for calibrating the field of view Mount the EVOS Calibration Slide supplied EVOS FL Auto Imaging System in the vessel holder as shown in the wizard Choose an objective from the options available and click Next Click Adjust Lighting to select the Monochrome camera and adjust the light and camera settings Click Adjust Focus to focus on the calibration slide Click Center Crosshair to match the crosshairs on the screen to the crosshairs on the calibration slide Once you have matched the crosshairs click Next Use the Auto button to get the lines close to the circle i e calibration target on the screen Using the directional arrows move the lines so that they touch just the outside edges of the circle Click Next and repeat the procedure for the Color camera After completing the calibration procedure click Finish Mount the slide as shown Choose an objective AMG 60X Plan FL AMEP 4626 AMG 4X Plan PH AMEP 4632 AMG 10X Plan FL AMEP 4623 AMG 20X Plan FL AMEP 4624 AMG 40X Plan FL AMEP 4625 lt Back Next gt Cancel 1 Adjust Lighting 2 Adjust Focus 3 Match the crosshairs on the sc Adjust Lighting Adjust Focus Center Crosshair Cancel 1 Use the Auto button to get the lines close 2 Move the lines so that they
22. unscrew and remove the vessel holder adaptor before removing the base plate Incubator Master Plate X Y stage base plate Vessel holder adaptor tif A X Y stage i EVOS FL Auto 2 Secure the onstage incubator master plate to the X Y stage using the four thumb screws indicated by red arrows Onstage incubator master plate X Y stage b V0s FL Auto 51 Assemble the Stagetop Environmental Chamber D2 1 Place the incubator chamber on the onstage incubator master plate and secure it in place using the four 2 0 mm hex screws indicated by red arrows Incubator chamber gt Onstage incubator master plate 2 Attach the vessel holder adaptor to the incubator chamber using the four thumb screws indicated by red arrows Vessel holder LS Note Place an empty dummy culture plate into the vessel holder adaptor forthe initial warm up and equilibration to prevent build up of condensation on the optical components and the inside of the EVOS FL Auto Imaging System 3 Place the heated glass lid with the no fog glass window on the incubator chamber The heated glass lid is guided and held secure in its place by the two magnets on its rim Heated glass lid 4 Place the light shield with tinted plastic window on top of the heated glass lid Use of the light shield is required for fluorescence imaging applications 5 If desired place the light shield cover on the
23. 06 Using the EVOS Onstage Incubator Turn ON the EVOS Onstage Incubator Configure Gas Connections 1 Turn ON the EVOS FL Auto Imaging IMAGE scan IMEWABSE system System as described on page 6 Re ON review Turn the computer and monitor ON Environment Chamber Setup Turn ON the power switch to the Control Unit v Enable Environment Chamber Restart the EVOS FL Auto software Go to the Time Lapse tab The Time Lapse tab will now display the one Incubator tab Humidity 3 Click Enable Onstage Incubator under coz so Time Lapse gt Incubator tab oxygen 0 0 Note Place an empty dummy culture plate into the vessel holder adaptor fortheinitial warm up and equilibration to prevent build up of condensation on the optical components and the inside of the EVOS FL Auto Imaging System 1 Goto System gt Basic tab page 30 and Configure Gas Connections then click Configure Gas Connections ort No Connection 2 Select the appropriate option for each E ee 5 Premixed CO2 Port that reflects your set up for the EVOS Onstage Incubator For Port 1 you may choose 5 pre mixed CO or air or manually enter the percentage of the CO and oxygen to reflect the specifics of your set up Port 2 is reserved for Nitrogen and Port 3 is reserved for CO only Choose No Connection if you are not using that particular port in your set up 3 Click OK once you have configured the gas connections for
24. Giear An Channels Apply Pseudo Color allows the a display of the images in user defined pseudo color see page 22 when using the Monochrome camera in fluorescence channels amp Note While pseudo colors help differentiate the channels used in multi channel overlays grayscale images usually show more detail Actual Mode allows you to individually adjust the LED intensity gain and exposure time rather than controlling them as a single parameter with the Light Intensity slider Clear All Channels clears out all memory buffers LS Note Memory buffers are temporary holding areas for captured images Channel Buttons Channel buttons are used for selecting the desired channel from the number of installed LED light cubes or the transmitted light from the condenser The selected channel is indicated by the blue color of the corresponding Channel button Note If there are images in the memory buffers they will be displayed as thumbnails below the corresponding light cube indicators Snap to Last Button Quick Slider Find Sample and Auto Fine Buttons Coarse and Fine Focus Sliders Hot Buttons Capture Image Capturing Images using the Capture Tab Snap to Last button reverts to the focus position of the last captured image Quick slider moves the focus through a small subset of the total focus range determined by the current vessel Utilizing the Quick slider drastically elim
25. R CHANNEL SAVE IMAGE 9 Click Save Image to save the images to the desired location For more information see page 13 Capture Capture Tab Controls Clicking Capture on the Home page page 7 or selecting the Image Capture tab i e the Capture tab under Image on any screen provides access to Capture Controls Capture Controls can also be accessed using the Quick Image Wizard see Quick Image page 8 IMAGE SCAN TIME LAPSE SYSTEM a te a ii iu E S O G z 17 w 8 2 6 l F FIND SAMPLE AUTO FINE F CAFE cr or 10 D 12 6 Vessel Expert button Snap to Last button Capture button Virtual vessel Quick focus slider Capture All button Objective buttons Find Sample button Z stack button Power toggle Auto Fine button Save button Light and Camera button Coarse focus slider Light intensity slider Channel buttons Fine focus slider X Note and are Hot Buttons Hot Buttons are user specified programmable buttons that customize the imaging system for your specific needs For more information on customizing Hot Buttons see page 21 Vessel Expert Button Virtual Vessel Objective Buttons Power Toggle Light and Camera Button Vessel Selection Wizard Vessel Expert button opens the Vessel Selection Wizard which is used for elect Ceres selecting the sample vessel category such as well p
26. Sensor data cable Stagetop with 6 pin connector environmental chamber ee j J is 7 TRE EO o Life RER AMC1000 EN F1713 1814 006 ee eal 9 cr Jane 2913 A eg x a Sensor data cable jack USB control Sen cable jack Power input jack Port 1 Air Port 2 N Port 3 CO 49 EVOS Onstage Incubator Stagetop Environmental Chamber Light shield cover Light shield Heated glass lid Vessel holder adaptor Incubator chamber Master plate 00 The Stagetop Environmental Chamber of the EVOS Onstage Incubator consists of the incubator chamber the vessel holder adaptor the heated glass lid the light shield and the light shield cover The Stagetop Environmental Chamber sits on the onstage incubator master plate attached to the X Y stage of the EVOS FL Auto Imaging System see Assemble the Stagetop Environmental Chamber page 52 Stagetop environmental chamber assembled Installing the EVOS Onstage Incubator Before setting up the EVOS Onstage Incubator unpack the unit and accessories and verify all parts are present Refer to page 47 for a list of the standard items included with the EVOS Onstage Incubator Contact your local distributor or Technical Support see page 71 if anything is missing Install the Onstage 1 Remove the X Y stage base plate from the X Y stage by unscrewing the four 3 0 mm screws on the base plate indicated by red arrows If necessary
27. Slide 2 Click Add Beacon and then click on the live image on the screen to add your beacons SELECTED WNone e Beacon 1 determines the top left starting position of the scan e Beacon 2 determines the bottom right of the scan When prompted adjust the focus and lighting for each of the channels that you have selected Determine how often you would like the system to refocus during a scan by selecting Find Sample and Auto Fine options for the scan e Find Sample searches over a wide range to provide coarse focus e Auto Fine searches over a narrow range to provide fine focus e You may choose to run either both or neither of the two focus functions during a scan routine Proceed to Finish Creating a Scan Routine page 26 29 Finish Creating a 1 Select how the images should be saved Enter a file name and click Browse to choose a location in the network or the USB drive Click Next to select the routine name and location Scan Routine lt Back Next gt Cancel How should the images be saved File name Folder IUSB1 EVOS Browse 2 Enter the name for the scan routine you have created and select the desired location to save the routine 3 Finish creating your routine by clicking Save to save your routine in the selected location without running it 4 Alternatively finish by clicking Save and Run and then Start to save your routine and automatically run your new routine Save an
28. USER GUIDE EVOS FL Auto Imaging System for Fluorescence and Transmitted Light Applications Catalog Numbers AMAFD 1000 Publication Number MANO007986 Revision 3 0 9 For Research Use Only Not for use in diagnostic procedures technologies Information in this document Is subject to change without notice DISCLAIMER LIFE TECHNOLOGIES CORPORATION AND OR ITS AFFILIATE S DISCLAIM ALL WARRANTIES WITH RESPECT TO THIS DOCUMENT EXPRESSED OR IMPLIED INCLUDING BUT NOT LIMITED TO THOSE OF MERCHANTABILITY FITNESS FOR A PARTICULAR PURPOSE OR NON INFRINGEMENT TO THE EXTENT ALLOWED BY LAW IN NO EVENT SHALL LIFE TECHNOLOGIES AND OR ITS AFFILIATE S BE LIABLE WHETHER IN CONTRACT TORT WARRANTY OR UNDER ANY STATUTE OR ON ANY OTHER BASIS FOR SPECIAL INCIDENTAL INDIRECT PUNITIVE MULTIPLE OR CONSEQUENTIAL DAMAGES IN CONNECTION WITH OR ARISING FROM THIS DOCUMENT INCLUDING BUT NOT LIMITED TO THE USE THEREOF NOTICE TO PURCHASER LIMITED USE LABEL LICENSE Research Use Only The purchase of this product conveys to the purchaser the limited non transferable right to use the purchased amount of the product only to perform internal research for the sole benefit of the purchaser No right to resell this product or any of its components is conveyed expressly by implication or by estoppel This product is for internal research purposes only and is not for use in commercial applications of any kind including without limitation quality cont
29. a variety of advanced imaging and analysis tools All images acquired can be saved in JPEG BMP TIFF and PNG formats Key features of the EVOS FL Auto software include Quick Image Wizard Provides step by step instructions for capturing perfect images ideal for even the first time users Manual Exploration Allows control over every aspect of the system through a simple user interface using either touch screen monitor or mouse Routines Allows the creation saving and running of user defined routines to automate image collection Autofocus Can be setup in several different modes to optimize speed and accuracy Image Stitching Allows the scanning of an area to acquire multiple images to build a tiled and stitched image The Scan Review tool allows zooming in out and panning of the composite image The entire scan or only regions of interest can be exported Z Stacking Captures a series of images in the z axis The Z Stack Flat Focus feature collects a series of images extracts the most in focus pixels from each image and then returns a single focused image even if the sample is very thick Time Lapse Creates and runs time lapse movies based on user specifications Image Editing Provides built in image analysis tools in the user interface including the Manual Counting Tool Measure and Annotate and Image Review The EVOS FL Auto system is designed to be used for a broad range of applications including but not li
30. ains the relevant hazard see Safety Labels on Instruments These safety symbols may also appear next to DANGERS WARNINGS and CAUTIONS that occur in the text of this and other product support documents Symbol PEPEE LS gt Description Indicates that you should consult the manual for further information and to proceed with appropriate caution Indicates the presence of an electrical shock hazard and to proceed with appropriate caution Indicates the presence of a hot surface or other high temperature hazard and to proceed with appropriate caution Indicates the presence of a laser inside the instrument and to proceed with appropriate caution Indicates the presence of moving parts and to proceed with appropriate caution Indicates the presence of a biological hazard and to proceed with appropriate caution Indicates the presence of an ultraviolet light and to proceed with appropriate caution The following symbol applies to all Life Technologies electrical and electronic products placed on the European market after August 13 2005 Symbol Description Do not dispose of this product as unsorted municipal waste Follow local municipal waste ordinances for proper disposal provisions to reduce the environmental impact of waste electrical and electronic equipment WEEE European Union customers Call your Customer Service representative for equipment pick up and recycling See www lifetechnologies com for a l
31. al EVOS distributor for information Objective Lens Care Clean each objective periodically or when necessary with an optical grade swab and a pre moistened lens wipe or lens paper moistened with lens cleaning solution To avoid scratching the soft lens coatings use only optical grade cleaning materials and do not rub the lens Note To protect all optical components of the instrument use the dust cover when the instrument is not in use 33 Stage Care Clean the X Y stage as needed with paper towels or Kimwipes tissues dampened with 70 ethanol When moving the EVOS FL Auto Imaging System be sure to lock the X Y stage using the Shipping Restraint as shown on page 40 to prevent the stage from sliding Sterilization Procedures 34 Q IMPORTANT The EVOS FL Auto Imaging System should not be subjected to UV sterilization UV degrades many materials including plastic Damage from UV exposure is not covered under the manufacturer s warranty In case hazardous material is spilt onto or into the components of the EVOS FL Auto Imaging System follow the sterilization procedure as described below 1 2 Turn power OFF Clean the LCD display a Use a soft dry lint free cloth to wipe off any dust from the screen b Clean the LCD display with a non alcohol based cleaner made for flat panel displays IMPORTANT Do not spray cleaning fluid directly onto the screen as it may drip into th
32. andling potentially infectious materials Additional information about biohazard guidelines is available at www cdc gov In the EU Check your local guidelines and legislation on biohazard and biosafety precaution and the best practices published in the World Health Organisation WHO Laboratory Biosafety Manual third edition www who int csr resources publications biosafety WHO_CDS_CSR_LYO_ 2004_11 en 69 Safety and Electromagnetic Compatibility EMC Standards U S and Canadian Safety Standards European Safety and EMC Standards CE Australian EMC standards 70 This section provides information on e U S and Canadian safety standards e European safety and EMC standards e Australian EMC standards The CSA C US Mark signifies that the product meets applicable U S and Canadian standards including those from CSA CSA America ANSI ASME ASSE ASTM NSF and UL The CE Mark symbolizes that the product conforms to all applicable European Community provisions for which this marking is required Operation of the instrument is subject to the conditions described in this manual The protection provided by the instrument may be impaired if the instrument is used in a manner not specified by Life Technologies The C Tick Mark indicates conformity with Australian and New Zealand standards for electromagnetic compatibility Documentation and Support Obtaining Support Technical Support Safety Data She
33. d Run Run the scan File name Stored Routines Images 24 Slides 1 Folder USB1 EVOS Browse Elapsed time 00 00 00 Time remaining 00 00 00 Disk Space 50M TIP To run this routine press Save select the routine and press Run Recall a Saved Scan 1 On the Routines tab page 23 click Browse to find desired routine Routine 2 Select the routine from Stored Routines 3 Click Run to run the selected routine Click Edit to make changes to the routine parameters Click Delete to delete the routine 26 Editing Scans Exportings Scans Reviewing Scans Use the Scan gt Edit tabs Overlay Measure and Count to edit your scans The controls for editing scans are identical to the controls used for editing single captured images in Image gt Edit tabs Refer to Editing and Analyzing Images on page 14 for detailed information on using these tools Scans are saved as sti files These files are very large and make exporting and opening them on other machines very difficult To make scans user friendly you have the option of exporting the scan using a different image quality You may also use the digital zoom to select a specific area of the image and then export that particular part of the screen To export a scan 1 Goto Scan gt Edit tab 2 In Save Settings select the desired Quality Low Medium or High The export tool will display the resolution and approximate size of the selected
34. d by red arrow in the example using the Position and Size controls v Slide 2 Position Which part of the slide would you like to scan Slide 1 Determine how often you would like the system to refocus during a scan by selecting Find Sample and Auto Fine options for the scan e Find Sample searches over a wide range to provide coarse focus e Auto Fine searches over a narrow range to provide fine focus e You may choose to run either both or neither of the two focus functions during a scan routine Scan Routine Options Placing Beacons on Live Image Select the amount of coverage You may choose from the available coverage options full coverage four points etc select to enter a number of random images enter a percentage of the vessel to scan or create a custom coverage erid A larger area requires longer scan time and results in a bigger file size Full coverage Four points Random Outer region only o Center region only Grid When prompted adjust focus and lighting for each of the channels that you have selected Proceed to Finish Creating a Scan Routine page 26 AREA 1 Place beacons at opposite corners of scan region When prompted choose Place beacon on the live microscope image MODE Pan Add Beacon Select Beacon A A Select the Pan mode and using the crosshairs on the virtual vessel pan to your area of interest in example on the right center of
35. d in Life Technologies document appears with an open triangle figure that contains a hazard symbol These hazard symbols are identical to the hazard icons that are affixed to Life Technologies instruments see Safety Symbols Symbols on Instruments Electrical Symbols The following table describes the electrical symbols that may be displayed on Life on Instruments Technologies instruments Description Indicates the On position of the main power switch Indicates the Off position of the main power switch Indicates a standby switch by which the instrument is switched on to the Standby condition Hazardous voltage may be present if this switch is on standby Indicates the On Off position of a push push main power switch Indicates a terminal that may be connected to the signal ground reference of another instrument This is not a protected ground terminal Indicates a protective grounding terminal that must be connected to earth ground before any other electrical connections are made to the instrument Indicates a terminal that can receive or supply alternating current or voltage Indicates a terminal that can receive or supply alternating or direct current or voltage 61 Safety Symbols Environmental Symbols on Instruments 62 The following table describes the safety symbols that may be displayed on Life Technologies instruments Each symbol may appear by itself or in combination with text that expl
36. desired imaging options such as vessel type area to capture lighting magnification etc and adjust the illumination and the focus Click the desired Capture button to acquire your image 5 Click Save Image to save the image to the desired location For more information see page 13 Save Save Tab Controls Clicking the Save Image button on any screen opens the Image gt Save tab which allows you to name and save an image file or a scan in a desired location in a USB drive or the network and to choose the desired Save Options SAVE IMAGE Folder address bar Describes the IMAGE SCAN TIME LAPSE SYSTEM location of the current folder or the file oC lees oe EY Up Folder button Navigates up to a ff higher level folder Felder USB1 Images Current folder Contains lower lever folders and or saved files Click on the lower level folders to navigate there Create Folder button Creates a new folder in the current location File Name text field Allows you to enter a file name for your image Save Options Allows you to save is your image using the Default File Name BPAE_Bodipy MitoTrackerRed_NucBlue tif Quantitative Monochrome or Custom trans options Bit Depth Allows you to set you color depth to 8 bit 256 colors or 16 bit thousands of colors high color File Type drop down menu Allows you choose the format of your saved image file Available options are JPEG
37. ductor disconnected DANGER ELECTRICAL HAZARD Use properly configured and approved line cords for the voltage supply in your facility gt gt e D DANGER ELECTRICAL HAZARD Plug the system into a properly grounded receptacle with adequate current capacity The EVOS FL Auto Imaging System has an installation overvoltage category of II and is classified as portable equipment Physical Hazard Safety Moving Parts WARNING PHYSICAL INJURY HAZARD Moving parts can crush and cut Keep hands clear of moving parts while operating the instrument Disconnect power before servicing the instrument Biological Hazard Safety a WARNING BIOHAZARD Biological samples such as tissues body fluids and blood of humans and other animals have the potential to transmit infectious diseases Follow all applicable local state provincial and or national regulations Wear appropriate protective eyewear clothing and gloves Read and follow the guidelines in these publications In the U S U S Department of Health and Human Services guidelines published in Biosafety in Microbiological and Biomedical Laboratories stock no 017 040 00547 4 www cdc gov OD ohs biosfty bmbl4 bmbl toc htm Occupational Safety and Health Standards Bloodborne Pathogens 29 CFR 1910 1030 www access gpo gov nara cfr waisidx_01 29cfr1910a_01 html Your company s institution s Biosafety Program protocols for working with h
38. e display Lightly wipe working surfaces of the EVOS FL Auto Imaging System stage top objective turret housing etc with paper towels or Kimwipes tissues dampened with 70 ethanol or 4 000 ppm hydrogen peroxide H O IMPORTANT Do not allow sterilization solution to get into lubricated areas such as the stage roller bearings or any points of rotation such as stage motors condenser wheel etc Do not soak any surface in sterilization solution NEVER spray liquid anywhere on the EVOS FL Auto Imaging System Always wipe surfaces with dampened paper towels instead Software Updates Download Software Update Install Software Update Periodically Life Technologies adds functionality and other improvements to the EVOS user interface We recommend keeping your EVOS FL Auto Imaging System up to date with the latest EVOS FL Auto software If you have any questions about software updates contact your local EVOS distributor If you do not have your distributor information contact Technical Support page 71 Software updates are available from the EVOS FL Auto Imaging System product page at www lifetechnologies com evosflauto 1 Download the update directly to the top level of a USB flash drive with at least 50 MB available Do not open or rename the file on your computer EVOS FL Auto Imaging System will verify and install it during the update process Download the current user guide for the EVOS FL
39. ets SDS Limited Product Warranty For the latest services and support information for all locations go to www lifetechnologies com At the website you can e Access worldwide telephone and fax numbers to contact Technical Support and Sales facilities e Search through frequently asked questions FAQs e Submit a question directly to Technical Support techsupport lifetech com e Search for user documents SDSs vector maps and sequences application notes formulations handbooks certificates of analysis citations and other product support documents e Obtain information about customer training e Download software updates and patches Safety Data Sheets SDSs are available at www lifetechnologies com sds IMPORTANT For the SDSs of chemicals not distributed by Life Technologies contact the chemical manufacturer Life Technologies Corporation and or its affiliate s warrant their products as set forth in the Life Technologies General Terms and Conditions of Sale found on Life Technologies website at www lifetechnologies com termsandconditions If you have any questions please contact Life Technologies at www lifetechnologies com support 71 Headquarters 8 5791 Van Allen Way Carlsbad CA 92008 USA Phone 1 760 603 7200 Toll Free in USA 800 955 6288 For support visit lifetechnologies com support or email techsupport dlifetech com technologies lifetechnologies com 8 July 2013
40. fe shipment of system see page 40 5 Care and Maintenance General Care e When cleaning optical elements use only optical grade materials to avoid scratching soft lens coatings e Use the appropriate cleaning solutions for each component as indicated in the Sterilization Procedures below e If liquid spills on the instrument turn off the power immediately and wipe dry e Do not exchange objectives between instruments unless you know that the components have been approved and recommended by Life Technologies e After using cover the instrument with the supplied dust cover Note Always use the correct power supply The power adaptor specifications appear on the serial number label front of LCD hinge and in the Specifications Damage due to an incompatible power adaptor is not covered by warranty A N CAUTION Never disassemble or service the instrument yourself Do not remove any covers or parts that require the use of a tool to obtain access to moving parts Operators must be trained before being allowed to perform the hazardous operation Unauthorized repairs may damage the instrument or alter its functionality which may void your warranty Contact your local EVOS distributor to arrange for service Q IMPORTANT If you have any doubt about the compatibility of decontamination or cleaning agents with parts of the equipment or with material contained in it contact Technical Support page 71 or your loc
41. g on the weight of the computer and or the monitor moving them may require two or more people Things to consider before lifting the computer and or the monitor e Make sure that you have a secure comfortable grip on the computer or the monitor when lifting e Make sure that the path from where the object is to where it is being moved is clear of obstructions e Do not lift an object and twist your torso at the same time e Keep your spine in a good neutral position while lifting with your legs e Participants should coordinate lift and move intentions with each other before lifting and carrying e Instead of lifting the object from the packing box carefully tilt the box on its side and hold it stationary while someone slides the contents out of the box Ensure that anyone who operates the instrument has e Received instructions in both general safety practices for laboratories and specific safety practices for the instrument e Read and understood all applicable Safety Data Sheets SDSs See Safety Data Sheets SDS CAUTION Using cleaning or decontamination methods other than those recommended by the manufacturer may compromise the safety or quality of the instrument ZN CAUTION PHYSICAL INJURY HAZARD The instrument is to be serviced only by trained personnel or vendor specified in the user guide Do not remove any covers or parts that require the use of a tool to obtain access to moving parts Opera
42. g the refresh a progress icon will appear e Unless there is an issue with the network or you are using an incompatible adaptor refreshing the connection should resolve the problem within a few moments Contact your network administrator for help if the problem persists e If your configuration requires using a static IP address select the Static IP Address option enter your IP Address Subnet Mask Gateway Address and DNS Server Address and then click Connect 31 Service Tab 32 Service tab contains information about the EVOS FL Auto Imaging System as well as several user preferences and settings Using the Service tab you may view and or change the following This EVOS Auto displays the Serial Es SCAN TIMELAPSE SYSTEM Number specific to each instrument and sasic menwonk i Software Revision which is the current CONTACT LIFE TECHNOLOGIES FOR SERVICE version of the software running on the This EVOS Auto system Ce O Update Software Use the button to DR A io tee eee Software Vessels sie upload new software versions from a USB flash drive see page 35 fers Microscope Position Obiective Lens Import Vessels Allows you to add BPI AMG 60x Plan FL AMEP 4625 AMG 4X Plan PH AMEP 4632 x AMG 10X Plan FL AMEP 4623 2 3 5 additional vessel types to the Vessel Selection Wizard Error Log Used for capturing error logs when prompted by the software Plug
43. ger life span gt 50 000 hours versus 300 hours for a typical mercury bulb and 1 500 hours for a metal halide bulb Each user interchangable auto configured light cube contains an LED collimating optics and filters In addition to the channel dedicated to the transmitted light from the condenser for brightfield contrast applications the EVOS FL Auto Imaging System can accommodate up to four fluorescent or specialty light cubes for multiple fluorescence research applications The table below lists some of the common fluorescent and specialty light cubes available from Light Technologies For a complete list of available light cubes and to inquire about custom light cubes go to www lifetechnologies com evosflauto or contact Technical Support see page 71 Light cube Dye DAPI DAPI Hoechst BFP TagBFP TagBFP CFP ECFP Lucifer Yellow Evans Blue GFP GFP Alexa Fluor 488 SYBR Green FITC YFP EYFP acridine orange DNA REP RFP Alexa Fluor 546 Alexa Fluor 555 Alexa Fluor 568 Cy 3 MitoTracker Orange Rhodamine Red DsRed Texas Red Texas Red Alexa Fluor 568 Alexa Fluor 594 MitoTracker Red mCherry Cy 3 5 Cy5 Cy 5 Alexa Fluor 647 Alexa Fluor 660 DRAQ5 Cy5 5 Cy 5 5 Alexa Fluor 660 Alexa Fluor 680 Alexa Fluor 700 Cy7 Cy 7 IRDye 800CW Specialty light cubes Dye CFP YFP em CFP YFP for FRET applications AO Acridine orange RNA simul
44. h the gas lines to the Control Unit via the PTC connectors for the appropriate gas intake port e If using pre mixed air attach to Port 1 Air In e If using compressed air and CO attach to Port 1 Air In and Port 3 CO In e For oxygen displacement attach to Port 1 Air In and Port 2 Nitrogen In 5 Plug the 6 pin sensor data cable to the Stagetop Environmental Chamber and the appropriate input jack on the Control Unit 6 Assemble the water reservoir and add warm water to the max fill line through the fill hole see image below Do not overfill the water reservoir warm water 7 Place the water reservoir into the Control Unit with the fill holes to the front and close the lid 8 Attach the heated hose between the Stagetop Environmental Chamber and the Control Unit 9 Plug the hose heater cable to the connector on the heated hose 99 Install the Light Before operating the EVOS Onstage Incubator the light shield retainer see image Shield Retainer below should be used to secure the incubator to the X Y stage to restrict the user from removing the light shield during operation 1 Place the Stagetop Environmental Chamber onto the EVOS FL Auto Imaging System X Y stage 2 Slide the light shield retainer over the Stagetop Environmental Chamber Make sure the light shield cover is in place 3 Use the thumbscrews to tighten the light shield retainer into the onstage incubator master plate
45. hlight saturated pixels in red option in the Settings gt Basic tab page 30 41 Software Interface Issues We recommend keeping the EVOS FL Auto Imaging System up to date with the latest software For more information see Software Updates page 35 Problem Possible Solutions Image does not respond to changes in Fe Click the Power button to return to live observation focus or stage position Scalebar does not appear when clicked Capture image first scalebar is only available after capturing an image Save button does not respond when Click capture first It is only possible to save an image that is clicked captured e Verify physical cable connections confirm the Ethernet jack is active Unable to connect to network o e Contact your network administrator to resolve any network issues Mechanical Issues Problem Possible Solutions Automatic stage does not move Remove shipping restraint Vessel does not sit securely on moving Use the correct vessel holder for the application refer to stage www lifetechnologies com evosflauto 42 Appendix A Description of EVOS FL Auto Imaging System Technical Specifications Physical Characteristics Hardware Note Specifications of the EVOS FL Auto Imaging System are subject to change without notice For the latest product information see the EVOS FL Auto Imaging System product page www lifetechnologies com ev
46. ho ew ange tanec nuaa at aan aac te ea eenen nanos 15 CO er na eT ee ee ee ee 16 ADO COULISSES tienen eau Re A tices eats eas NON 18 HI AOC RENEW coe sx tes cesarean dd ant sees sae vote dd an ee ds et us TT Sesto 20 4s Advanced ODS QUO Sis en an nd A nn dt nn 21 AA SDS Re EE a OC re eo eee ones Don ot 21 1 D Rene E PPO Oe PPD PP era ee E 23 TE La of SR ee Een een 28 A D T A 30 5 Care and Maintenance cccccccceccseceeceeeeeeeeesaeeeeesaesaeesaesaeeeeesaeseeesaesaneseeseeaeesaes 33 COCA Bee oc Rd OP IEE EEES T en 33 Obj chve Lens Came SR een nt nes mec nn ete no eee onde aise 33 Stage C i TP den ee tue 34 SterimliZatiOn Procedures RSR a A Ant A dt AU de AU 34 DOLEW abe Upda leS sin nn RS das 35 Calibrating the EVOS FL Auto Imaging System 36 Chaine the V OS LED Cieht C UDe nue conne connut 39 Installing theonipping IRSC A te 40 D TOWLE SIO OUAIS SE ce a eta te ee tet ne nn tac 41 Moce AU ry LS esas a na nae ears aaa ie a ao do eee 41 DOLEWare Miter ace Soues ieir ee Le etai titine ats 42 VRC CMe IGa TS SU CS ed Dies en tiens etes hic eut ne 42 Appendix A Description of EVOS FL Auto Imaging System cceeecssseeeeeeeeeeeeees 43 Technical SO SCIELE ALONG Ra en a en do a 43 Mstrament Exterior Components srana RS Sn ds tant as ouens 45 Operation Prineples nd Technical Overview sn masters 46 Appendix B EVOS Onstage Incubator issus 47 Techical SP CCIM CATIONS ES SA T e NN 48 EVOS Onstage Incubator Exterior Components
47. ibed below 1 2 Turn power OFF Lightly wipe working surfaces of the EVOS Onstage Incubator with paper towels or Kimwipes tissues dampened with 70 ethanol or 4 000 ppm hydrogen peroxide H O Q IMPORTANT Do not soak any surface in sterilization solution NEVER spray liquid anywhere on the EVOS Onstage Incubator Always wipe surfaces with dampened paper towels instead D9 Appendix C Safety Safety Conventions Used in this Document Safety Alert Words 60 Four safety alert words appear in Life Technologies user documentation at points in the document where you need to be aware of relevant hazards Each alert word IMPORTANT CAUTION WARNING DANGER implies a particular level of observation or action Definitions 1 IMPORTANT Provides information that is necessary for proper instrument operation accurate installation or safe use of a chemical T CAUTION Indicates a potentially hazardous situation that if not avoided may result in minor or moderate injury It may also be used to alert against unsafe practices AX WARNING Indicates a potentially hazardous situation that if not avoided could result in death or serious injury T DANGER Indicates an imminently hazardous situation that if not avoided will result in death or serious injury This signal word is to be limited to the most extreme situations Except for IMPORTANT safety alerts each safety alert wor
48. inates the need to use the Find Sample button and the Coarse Focus slider Find Sample and Auto Fine Buttons are used for automatic coarse and fine focusing on the sample Coarse and Fine Focus Sliders are used for manually adjusting coarse and fine focus Hot Buttons are user specified programmable buttons that customize the imaging system Up to four Hot Buttons can be displayed on the Image gt Capture tab e Capture Captures the current image on the screen e Capture All Captures an image using each of the installed light cubes e Preset Utilizes the capture options selected in Quick Image Launch Wizard e Transfection Creates a two image overlay using one fluorescent image and one transmitted light image e Save Saves the captured images e Z Stack Captures a series of images along the z axis of the sample e For detailed information on customizing Hot Buttons to your specific needs see page 21 1 Place the vessel containing your sample on the stage using the appropriate vessel holder For the types of vessel holders available for use with the EVOS FL Auto Imaging System refer to www lifetechnologies com evosflauto or contact Technical Support page 71 2 Onthe Home page see page 7 click Capture The system will move to the Image gt Capture tab i e the Capture tab under Image allowing you to directly control the instrument to capture your images 3 Using the Capture Tab Controls select the
49. ish to revisit 2 When prompted select a Vessel from the list Selected vessel type will be highlighted in blue and a virtual vessel will appear under the list in the example below a dual slide holder is selected Single Slide Holder Well Plate 4 Well Plate 6 Well Plate 12 Well Plate TEST VESSEL 12 Well Plate 24 Well Plate 96 Well Plate 23 Scan Routine Options Using an Area Rectangle 24 3 Select objectives camera and light channels to use in your scan routine 60x 4x pus Monochrome Color Fluorescence Phase Brightfield Phase Choose the Area you would like to cover in your scan by selecting an area rectangle or by placing beacons on a live image e Select area rectangle Move and size an area of interest rectangle on your virtual vessel If using this method continue with Routine Options Using an Area Rectangle below e Place beacons Mark the opposite corners of your area of interest on the live image to designate your area of interest If using this method continue with Routine Options Placing Beacons on Live Image page 25 When prompted choose Select area rectangle Using the virtual vessel vessel image representing the sample vessel select the wells or slides to cover in your scan in the example below Slide 2 in a dual slide holder is selected Select the area of the vessel you wish to scan by moving and sizing the area rectangle indicate
50. isplays or hides the tags onscreen Show Grid Displays or hides the onscreen counting grid Grid size Selects the erid size from a drop down menu Save Image as Screenshot Enables the saving of the image showing the labels counts and percentages as displayed in the Count tool Deselecting this option produces an image saved with the tags only Save Image Saves the image in the desired location Aa Note You can document the results of your count by saving the tagged image with the Save Image as Screenshot option selected this will save the image showing the labels counts and percentages as displayed in the Count tool Count Procedure Begin by acquiring an image in the Capture tab see page 12 or by opening a saved image using the Review tab see page 20 Click Show Grid and choose Grid size from the drop down menu You may also choose to leave this option inactive Click the text field next to a label to enter a name for that label category Repeat for the remaining labels as necessary You can use up to six labels per count To begin counting click Add to activate the Add mode Select a Label button and then left click at each point onscreen to tag the items for that category Each tagged item will show the label used and a running tally of the count with the percentage for the label used and the total count will be displayed in the Count tool BS Note You may switch labels
51. ist of customer service offices in the European Union Safety Labels on Instruments The following CAUTION WARNING and DANGER statements may be displayed on Life Technologies instruments in combination with the safety symbols described in the preceding section Hazard Symbol English Francais JN CAUTION Hazardous chemicals Read the Safety Data Sheets SDSs before handling ATTENTION Produits chimiques dangereux Lire les fiches techniques de s ret de mat riels avant toute manipulation de produits CAUTION Hazardous waste Refer to SDS s and local regulations for handling and disposal ATTENTION D chets dangereux Lire les fiches techniques de s ret de mat riels et la r gulation locale associ es a la manipulation et l limination des d chets AN DANGER High voltage DANGER Haute tension WARNING To reduce the chance of electrical shock do not remove covers that require tool access No user serviceable parts are inside Refer servicing to Life Technologies qualified service personnel AVERTISSEMENT Pour viter les risques d lectrocution ne pas retirer les capots dont l ouverture n cessite l utilisation d outils L instrument ne contient aucune pi ce r parable par l utilisateur Toute intervention doit tre effectu e par le personnel de service qualifi venant de chez Life Technologies DANGER Class 3B visible and or invisible laser radiation presen
52. late Petri dish culture flask chamber slide etc and vessel type such as i 24 well plate 35 mm Petri dish T75 flask etc from the Category and Type drop down menus Well Plate WA Category With appropriate vessel holders the EVOS FL Auto Imaging System can accommodate a wide variety of common culture vessels types in standard sizes Note that a vessel MUST be selected prior to imaging LS Note For a list of the vessel holders and stage plates available for the EVOS FL Auto Imaging System refer to www lifetechnologies com evosflauto or contact Technical Support page 71 Virtual vessel is a graphical representation of the sample vessel as selected with the Vessel Expert To choose the location on the sample vessel you wish to view and image move the crosshairs to the corresponding location on the virtual vessel on the Image gt Capture tab Objective buttons control the active objective turret position The magnifications displayed on the Image Capture tab reflect the objective profiles chosen on the System Service tab Power toggle turns the LED light source in the selected channel ON and OFF Light and Camera button opens the Light and Camera Light and Camera selection tool for ee selecting the Monochrome camera am Fluorescence Phase Brighttieid Phase for fluorescence and brightfield or the Color camera for brightfield only Apply Pseudo Color
53. light shield for fluorescence imaging applications The light shield cover completely blocks any ambient light from entering the environment chamber and improves image quality in fluorescence imaging applications Light shield cover 6 When you are ready for acquiring images secure the Stagetop Environmental Chamber to the X Y stage using the light shield retainer see page 56 03 Set Up for Operation 04 Follow the procedure below to set up the EVOS Onstage Incubator for operation For the locations of the various input jacks and gas ports refer to EVOS Onstage Incubator Control Unit Instrument Panel page 49 Q IMPORTANT Do not position the Control Unit so that it is difficult to turn off the main power switch see page 49 for the location of the power switch In case of an instrument malfunction turn the main power switch to the OFF position and disconnect the power cord from the wall outlet 1 Plug power cord into the power input jack on the Control Unit and the wall outlet 2 Plug USB cable into the USB control cable jack on the Control Unit and the USB port on the EVOS FL Auto computer 3 Connect each gas line to the appropriate gas tank via the PTC push to click connectors threaded into the regulator To do this push the tubing into the PTC connector until it clicks into place Pull on tubing slightly to ensure a tight connection the tubing should not come out pue te ar 4 Attac
54. ly locating saved files It also allows you to review still images or play video files from the USB drive or the network connection You may also use this tab to rename or delete saved files 29 System Basic Tab The System gt Basic tab allows you to select basic system functions Live Expand Pinch Gesture canbe selected based on user preference Clicking Show Scale bar option displays the scale bar on captured IMAGE SCAN TIME LAPSE SYSTEM cea neron secs Live Expand Pinch Gesture Changes Objective images l Changes Zoom _ constrained Clicking Reset Scale Bar returns the Changes Zoom scale bar to its original location on the Jens screen Show Getting Started Button displays the shortcut button to the Home screen on the user interface amp Show Scale Bar on captured images Reset Scalebar location ba Show Getting Started Button Capture focus nominal is used to ee ii calibrate the Z range i e depth of vessels that are not yet pre configured within the auto user interface or to fine eenen tune the focus plane used by system for Find Sample and Auto Fine focus functions Calibrate Vessel Alignment Calibrate Vessel Alignment opens the Vessel Alignment Tool used for adjusting the alignment positions for the current vessel see page 38 Calibrate Oxygen Sensor is used for calibrating the oxygen sensor for the proper operation of the EVOS Onstage
55. mited to multi channel fluorescence imaging cell density assays multiple position vessel scanning and time lapse imaging Instrument Exterior Components Condenser Condenser slider slot contains automatic phase annulus selector USB port for PC connection Power ef input jack o i TR Power High resolution switch touch screen monitor Automatic X Y axis stage Lifting handholds for safe transport N Note The EVOS FL Auto Imaging System is a fully automated imaging system no manual adjustment is required for the objective focusing controls light cube and camera selection etc 45 Operation Principles and Technical Overview LED Illumination LED Light Cubes 46 The EVOS FL Auto Imaging System utilizes an adjustable intensity LED light source provided by the proprietary user interchangeable LED light cube see below Because the LED light source is as close as possible to the objective turret the number of optical elements in the channel is minimized High intensity illumination over a short channel increases the efficiency of fluorophore excitation providing better detection of weak fluorescent signals In contrast to traditional fluorescence microscopy light sources that use mercury a toxic carcinogen requiring special handling and disposal the LED light source of the EVOS FL Auto Imaging System is more environmentally friendly energy efficient and has a significantly lon
56. nd one transmitted light image e Save Saves the captured image with a single click v Highlight saturated pixels in red e Z Stack Captures a series of images along the z axis of the sample O To select the Hot Buttons you wish to display on the Image gt Capture tab 1 Click Mono or Color to select the camera Monochrome or Color for which you wish to apply the Hot Buttons 2 Select up to four Hot Buttons from Button drop down menus in the order you want them displayed Button 1 4 CAPTURE Button 1 Button 2 Button 3 co Z STACK Button 4 21 Selecting Pseudo Colors Highlight Saturated Pixels in Red 22 When using the Monochrome camera in multi channel overlays applying a pseudo color to each channel helps differentiate the images To assign a custom pseudo color to a channel follow the procedure below 1 In Image gt Settings select Mono for the Monochrome camera 2 Click Edit for the desired channel and select the color you wish assign 3 Repeat for the remaining channels Selecting Highlight Saturated Pixels in Red option displays pixels in saturated areas on a live image in red and gives you the opportunity to adjust the LED intensity gain and exposure time either individually in the Actual mode or together by using the Light intensity slider see page 11 Scan Routines Tab Controls Creating a Scan Routine The Scan tab gives you the op
57. nstrument box e Power cable e Power supply e Display Port to DVI cable e USB type A to B cable e White cardboard box contains the light box and vessel holders e Dust cover Confirm that the power switch is OFF located on the right side of the instrument base see page 45 Plug the power cable into the power supply and check for the light on the power supply Plug the power supply connector into the instrument Use the Display Port to DVI cable to connect the Display Port output on the computer to the DVI input on the monitor Use the USB cable to connect the instrument to the computer Note At this point everything should be plugged in and OFF Save the packaging for future shipping storage of the instrument D IMPORTANT Do not subject the EVOS FL Auto Imaging System to sudden impact or excessive vibration Handle the instrument with care to prevent damage Remove the The Shipping Restraint prevents the X Y stage from moving during shipping If the Shipping Restraint Stage is not secured during shipping shock and vibration can damage the motors that move the stage The Shipping Restraint must be removed before the EVOS FL Auto Imaging System is powered on Automatic X Y stage Shipping Restraint 1 Unscrew the three thumb screws until they spin freely You do not need to remove them from the Shipping Restraint altogether 2 Gently pull the Shipping Restraint forward away from the unit
58. oftware will use an algorithm to mark and count the regions of interets ROI on the screen see the example below The result of the count i e the number of ROI on the screen above the signal threshold is displayed in the Count field Review the ROI marked on the screen and adjust the Sensitivity of the count by moving the slider in the More or Less direction to include more or less ROI in the count To mark more regions in the the Threshold slider in the More or Less direction Ss Note Moving the slider in the More direction increases the count sensitivity _ by lowering the threshold above which the ROI are counted and results in a greater number of ROI and a higher count moving it in the Less direction increases the threshold value thereby reducing sensitivity and lowering the count 5 To save the image with the tags select Save Image As Screenshot and then click Save Image Image Review The Image gt Review tab utilizes a folder system for easily locating saved files and allows you to review still images or play video files from the USB drive or the network connection You may also use this tab to rename or delete saved files Review Tab Tools Folder address bar Describes the scan 1 de location of the current folder or the file Up Folder button Navigates up to a higher level folder Folder NETWORK3 EVOS P Current folder Contains lower lever folders and or saved files
59. option Save Settings Le TETILATE Resolution Approximate Size 364x273 Medium 910x682 3640x2730 29 81 MB 3 Click Export Scan to save it in the quality selected The Scan gt Review tab utilizes the same folder system as the Image gt Edit tab page 20 for easily locating saved files It also allows you to review still images or play video files from the USB drive or the network connection You may also use this tab to rename or delete saved files 27 Time Lapse Time Lapse Tab The Time Lapse tab gives you the option of creating and running time lapse routine to stitch together a video that shows changes over time in your samples Creating and The Time Lapse Wizard in the Time lt Back Next gt Cancel Running Time Lapse tab walks you through the steps Lapse Routines of creating a Time Lapse Routine to Time Lapse Wizard acquire individual images at given All scenes have been set up Please choose where you would like intervals over a time period based on TREST ONE ee REEE your specifications which can then be stitched together into a video Create video for each beacon 1 Click the Create New Routine v Include the following information as a watermark button to launch the Time Lapse Wend v Image v Time Lapse v Incubator Enter the Name of the routine v Save Individual Channels Select the objectives camera light channels and whether to use Autofocus option Z stack and Auto lighting PNG
60. or IMPORTANT safety alerts each safety alert word in a Life Technologies document appears with an open triangle figure that contains a hazard symbol These hazard symbols are identical to the hazard symbols that are affixed to Life Technologies instruments see Safety Symbols in Appendix C 1 Installation Standard Items Included Before setting up the EVOS FL Auto Imaging System unpack the unit and accessories and verify all parts are present Contact your distributor if anything is missing Note If you do not have your distributor information contact Technical Support see page 71 e EVOS FL Auto Imaging System e Computer e Touch screen monitor e Keyboard e Mouse e Light cubes as ordered e Objectives as ordered e Vessel holder s as ordered e Light box with cover e Light cube tool e Sliders Block slider Diffuser slider e Dustcover e USB flash drive includes User Guide and Quick Start Guide Operating Environment e Place the EVOS FL Auto Imaging System on a level surface away from vibrations from other pieces of equipment e Allow at least 5 cm 2 in free space at the back of the instrument to allow for proper ventilation and prevent overheating of electronic components e Set up the EVOS FL Auto Imaging System away from direct light sources such as windows Ambient room lighting can enter the imaging path and affect the image quality e Operating temperature range 4 32 C
61. ork automatically paso JS scrvice when it is connected to it via an Ethernet Address 10 1 104 235 Subnet 255 255 128 0 Gateway 10 1 1 1 cable the DHCP option is selected E and Connect is clicked Not connected to a network share If the network is not recognized r automatically do the following DHCP Static IP tt 1 Enter your network domain user name and password in Network Credentials fields and select a server to view the top level of shared folders on that server IP Address Subnet You may now navigate below the top ee level shared folders on the Network Domain Username page 2 After the server accepts your login the EVOS system will display the list of available shared folders on the selected server Select a shared folder and click Add ea a RE to include it in the list of possible file TEP Client Not Connected destinations You may also type in the file path and click Add 3 The folder should appear on the list box below the Add button If it does not contact your network administrator for help If you need to remove a shared folder from the destinations list select the folder name and click the Remove button e The EVOS FL Auto Imaging System will try to connect for about 30 seconds If there is a problem with the connection the Network page will display Not connected to a network share e Double check the physical connections and click the Refresh Network button Durin
62. osflauto Height 32 2 cm 12 7 in Depth 47 2 cm 18 6 in Width 34 3 cm 13 5 in Weight 20 0 kg 44 1 Ib Footprint Approximately 92 cm x 92 cm 36 in x 36 in entire system includes the instrument computer and 22 touch screen high resolution LCD monitor Operating temperature 4 32 C 40 90 F Operating humidity lt 90 non condensing Operating power 100 240 VAC 1 8 A Frequency 50 60 Hz Electrical input 24 VDC 5A Illumination Adjustable intensity LED gt 50 000 hour life per light cube Contrast methods Epifluoresence and transmitted light brightfield and phase contrast Objective turret 5 position automated control Fluorescence Channels Simultaneously accommodates up to 4 fluorescent light cubes Condenser working distance 60 mm Stage Automated X Y scanning stage travel range of 115 mm x 78 mm with sub micron resolution interchangeable vessel holders available LCD display 22 inch high resolution touch screen color monitor Camera Dual monochrome and color camera Monochrome high sensitivity interline CCD Color high sensitivity CMOS Output ports Multiple USB ports 1 display output with DVI adaptor supports direct output to USB and networked storage 43 EVOS FL Auto Software Applications 44 The integrated EVOS FL Auto software accessed by a touch screen monitor or the mouse features standard functions such as a scale bar and image review tool as well as
63. reviously saved image to perform Auto Count From File File 20130305_HeLa_H2BGFP_10X png Count v Separate touching cells O Count field displays the results of the Count 198 count i e the number of ROI on the screen above the signal threshold a Moro Separate touching cells option allows you to count cells that are touching seperately and increases the accuracy of the count Sensitivity slider controls the sensitivity of the Auto Count tool by changing the threshold of how the software perfoms auto counting e Move the slider in the More direction to increase auto count sensitivity i e decrease detection threshold to mark and count more ROI e Move the slider in the Less direction to decrease auto count sensitivity i e increase detection threshold to __ mark and count less ROI sde Save Image as Screenshot option enables the saving of the image showing the labels counts and percentages as displayed in the Count tool Deselecting this option produces an image saved with the tags only Save Image button saves the image in the desired location Auto Count Procedure Begin by acquiring an image using Capture see page 12 Go to the Auto Count tab and then click Count to initiate the Auto Count procedure Alternatively click Count From File to perform the Auto Count procedure on a previously saved image Based on a histogram of the captured image the EVOS FL Auto s
64. rol and commercial services such as reporting the results of purchaser s activities for a fee or other form of consideration For information on obtaining additional rights please contact outlicensing dlifetech com or Out Licensing Life Technologies Corporation 5791 Van Allen Way Carlsbad California 92008 TRADEMARKS The trademarks mentioned herein are the property of Life Technologies Corporation and or its affiliate s or their respective owners Cy is a registered trademark of GE Healthcare UK Limited DRAQ5 Is a registered trademark of Biostatus Limited Windows is a registered trademark of Microsoft Corporation Kimwipes is a registered trademark of Kimberly Clark Corporation ViewSonic is a registered trademark of ViewSonic Corporation 2013 Life Technologies Corporation All rights reserved Contents POOUL PAIS GUNG Re ne ea eo do nano no Ge di 3 Ms INSTALLATIONS Rd en dinde 4 DbaING at CI CO SCM CCl ss tice dito ann De de dant ti ne an nn ete one minis ts lee eee cite 4 OPE TALIS EMV IFONMENE SANS SA AE es act ii dustin Ne sa 4 D PIT nn 5 2 BASIC ODS Fa Ul OM En a dures nissan anse cts eee niet 8 CURSEUR RNA A oe Dalat Sed ea SA ppm Saat aaa sted ousted iataah dat 8 CADRE Rd een eines 10 Capiure MAR ES ea tetes cu de bte tendre 12 DAME Re D sales he csc A dl AA a A A 13 3 Editing and Ana ZING MAGES na nt ont 14 EEE EEE 14 IN LCS CITC tsps sos terse oe wena ng saab a heron ines beds ett de wba aaa we oraae eacaain
65. s Association CGA 580 cylinder adaptor on the inlet side and a Swagelok type end fitting that accepts 6 35 mm 0 25 in o d tubing The primary gauge 0 to 3000 psi 0 to 25 000 kPa recommended measures tank pressure and the secondary gauge 0 to 200 psi 0 to 2000 kPa recommended measures regulated pressure The secondary gauge must allow regulation to 50 psi Compressed Gas Association CGA 580 cylinder adaptor with a needle type shutoff valve on the exit side The needle valves should have Swagelok type end fittings ready for connection to 6 35 mm 0 25 in o d tubing Attaching the cylinder Attach the pressurized gas cylinder firmly to a wall or gas cylinder cart by means of approved straps or chains Ventilation WARNING The Onstage Incubator should be installed and operated in a requirements well ventilated environment as defined as having a minimum airflow of 6 10 air changes per hour Please contact your environmental health and safety coordinator to confirm that the Ion instruments will be installed and operated in an environment with sufficient ventilation Ventilation requirements Allow at least 50 cm 20 in of clearance around the Instrument for ventilation 65 Chemical Safety Chemical Hazard WARNING CHEMICAL HAZARD Before handling any chemicals refer Warning to the Safety Data Sheet SDS provided by the manufacturer and observe all relevant precautions Ak WARNING CHEMICAL HAZARD All
66. ser look at live or captured images To zoom on an image click Zoom and oe aspect nario then double click the area on the screen wien PTE you wish to enlarge To restore the view to unzoomed magnification right click anywhere on the image a Annotation Tools Include the text Ein oe box arrow histogram rectangle ellipse and line tools Save Image As Screenshot v Select Annotation after it is added If there is no keyboard connected to the system a virtual keyboard will appear when adding or editing a text box A SAVE IMAGE D Annotation Properties Edit Text Allows you to edit existing text in a selected text box Color Edit button Allows you to select the color of an annotation Measure Selects what is measured in a region of interest ROI Width Changes the line width of the selected annotation Misc Delete All Annotations Deletes all existing annotations Show Annotations Option to display or hide the annotations on screen Show Histogram Option to display or hide the histogram for a selected ROI Save Image as Screenshot Option for saving the image shown onscreen as its own file Units Selects the units of measurement used for the annotations Save Image Saves the image and the annotation s as a single file For more information see page 13 amp Note All annotations will remain whether or not a new image is captured To start over
67. spill occurs follow the manufacturer s cleanup procedures as recommended in the SDS e Comply with all local state provincial or national laws and regulations related to chemical storage handling and disposal 66 Chemical Waste Safety Chemical Waste Hazard Chemical Waste Safety Guidelines Waste Disposal CAUTION HAZARDOUS WASTE Refer to Safety Data Sheets SDSs and local regulations for handling and disposal To minimize the hazards of chemical waste Read and understand the Safety Data Sheets SDSs provided by the manufacturers of the chemicals in the waste container before you store handle or dispose of chemical waste Provide primary and secondary waste containers A primary waste container holds the immediate waste A secondary container contains spills or leaks from the primary container Both containers must be compatible with the waste material and meet federal state and local requirements for container storage Minimize contact with chemicals Wear appropriate personal protective equipment when handling chemicals for example safety glasses gloves or protective clothing For additional safety guidelines consult the SDS Minimize the inhalation of chemicals Do not leave chemical containers open Use only with adequate ventilation for example fume hood For additional safety guidelines consult the SDS Handle chemical wastes in a fume hood After emptying the waste container seal it
68. strument Q IMPORTANT Do not power the instrument back on until the Shipping Restraint has been removed 6 Troubleshooting Note For additional technical support contact your local EVOS distributor If you do not have your distributor information refer to www lifetechnologies com evosfauto or contact Technical Support page 71 Image Quality Issues Problem Possible Solutions Image is too dim at higher magnifications Remove condenser slider if one is in place Specks dots or blurs on image Follow instructions under Objective Lens Care page 33 to clean objectives Uneven focus across screen e Position specimen so that it lies flat on the stage ensure that the specimen s thickness is even e Be sure vessel holder is mounted flat with respect to stage Difficulty focusing on coverslipped specimen on standard slide Place the slide so the coverslip is facing up long working distance objectives require a thick optical substrate and image best through 1 0 1 5 mm of glass or plastic Image display is black e Click the Power button onscreen e Center specimen over objective e Verify power supply is connected and power switch is on Image display is red or red patches cover parts of the screen e Dim the illumination until the red highlights disappear to get the maximum level of brightness without any overexposed areas e Disable the Hig
69. t when open Avoid exposure to beam DANGER Rayonnement visible ou invisible d un faisceau laser de Classe 3B en cas d ouverture Evitez toute exposition au faisceau gt gt CAUTION Moving parts Crush pinch hazard ATTENTION Pieces en mouvement risque de pincement et ou d crasement 63 General Instrument Safety Moving and Lifting the Instrument Moving and Lifting Stand alone Computers and Monitors Operating the Instrument Cleaning or Decontaminating the Instrument Removing Covers or Parts of the Instrument 64 WARNING PHYSICAL INJURY HAZARD Use this product only as specified in this document Using this instrument in a manner not specified by Life Technologies may result in personal injury or damage to the instrument CAUTION PHYSICAL INJURY HAZARD The instrument is to be moved and positioned only by the personnel or vendor specified in the applicable site preparation guide If you decide to lift or move the instrument after it has been installed do not attempt to lift or move the instrument without the assistance of others the use of appropriate moving equipment and proper lifting techniques Improper lifting can cause painful and permanent back injury Depending on the weight moving or lifting an instrument may require two or more persons ZN WARNING Do not attempt to lift or move the computer or the monitor without the assistance of others Dependin
70. taneous green red with FL color AOred Acridine orange RNA CTC formazan Fura Red high Ca White Refracted light applications Appendix B EVOS Onstage Incubator EVOS Onstage Incubator Standard Items Included The EVOS Onstage Incubator is an optional accessory for the EVOS FL Auto Imaging System that enables the incubation of cells on the automatic X Y stage allowing the capture of images from the same sample over long periods of time and recording of time lapse movies The EVOS Onstage Incubator consists of a Stagetop Environmental Chamber that is placed on the automatic X Y stage of the imaging system and a separate Control Unit that supplies the power and gas air or air CO premix CO only and nitrogen only The onstage incubator is controlled by the same software and user interface that controls the EVOS FL Auto Imaging System e Stagetop Environmental Chamber e Control unit e Master Stage Plate also available separately as EVOS Onstage Master Plate Cat no MEPVH035 e Vessel holder for multi well plates also available separately as EVOS Onstage Vessel Holder Multiwell Plates Cat no AMEPVHO028 e Lid retainer shield e Cable with 6 pin connector e Cable USB A to B 180 cm 6 ft e Heated hose with temperature control 180 cm 6 ft also available separately as EVOS Onstage Incubator Hose Cat no AMEP4728 e Gas line 1 8 ID 1 4 OD also available separately as EVOS
71. tion to create and recall scan routines a series of steps to acquire a tiled and stitched image For repeat experiments Scan routines can be saved recalled even edited The Routines tab is used for creating and ss ee Ese SYSTEM running new scan routines and running Wo som son newew editing and deleting existing routines Create New Routine Guides you through a series of steps to create save and run a new scan routine based on your specifications Folder Describes the location of the current folder or routine Stored Routines Folder USB1 EVOS Browse Allows you to locate and select existing routines Stored list Contains lower lever folders and or saved routines Run Runs the selected routine Edit O Allows you to change the parameters of an existing routine Delete Deletes the selected routine 1 Goto Scan gt Routines tab and then click Create New Routine The EVOS software will guide you through the process of creating your own scan routine as you make a series of onscreen selections After making your selection at each step click Next for the next prompt or click Back to revisit the previous step Click Cancel to abort the process Back Next gt Cancel amp Note To have the option of changing a step of a saved scan routine any time you recall and run the scan select the Show this page again option at the bottom of the page you w
72. tors must be trained before being allowed to perform the hazardous operation Safety Requirements for EVOS Onstage Incubator WARNING Life Technologies recommends the use of nitrogen oxygen and carbon dioxide gas with the Onstage Incubator The use of alternative gasses is currently not supported and may adversely affect system performance Gas cylinders You must supply the required nitrogen oxygen and carbon dioxide gas cylinders and accessories for the installation This instrument requires pressurized house lines or one size 1 A gas cylinder that holds approximately 7 2 m 257 ft of gas when full for each gas Use only pre purified gasses of 99 9 or greater purity CAUTION Damage to the instrument and its products can result from using impure gas gases other than specified or an inadequate amount of gas WARNING EXPLOSION HAZARD Pressurized gas cylinders are potentially explosive Always cap the gas cylinder when it is not in use and attach it firmly to the wall or gas cylinder cart with approved brackets or chains A N WARNING Gas cylinders are heavy and may topple over potentially causing personal injury and tank damage Cylinders should be firmly secured to a wall or work surface Please contact your environmental health and safety coordinator for guidance on the proper installation of a gas cylinder Pressure regulator You must supply a two gauge regulator with a Compressed Ga
73. touch just the outside edges of the circle Bottom Objective AMG 40X Plan FL AMEP 4625 Diameter 0 2mm Status Next we will calibrate the Color Camera Calibrate Parfocality Parfocality means that the sample on the stage stays in focus when the objective is changed To judge whether the imaging system is properly set up and parfocality is properly adjusted the sample is focused at high power Then the objective is switched to a lower power and the refocused on the sample Because depth of field is greater at lower powers parfocality is always judged from high power to low A properly set up imaging system with correct parfocality will require very little refocusing The Parfocality Wizard of the EVOS FL Auto Software provides step by step instructions for easy calibration of parfocality to eliminate the need to refocus your camera s image before each image capture saving you time and frustration The process takes only a few minutes to complete 1 Inthe System gt Service tab page 32 click Calibrate Parfocality to open the Parfocality Wizard CERAR 2 Select the turret position on which to update the parfocality calibration by clicking the desired objective 2 Insert the parfocality grid slide supplied by Life Technologies so that button it resembles the graphic below 3 Mount the EVOS Parfocality Grid Slide supplied with the EVOS FL Auto Imaging System in the vessel holder as shown in the wizard 4
74. trument turn off the power immediately and wipe dry Clean the EVOS Onstage Incubator as needed with paper towels or Kimwipes tissues dampened with 70 ethanol Note Always use the correct power supply The power adaptor specifications appear on the serial number label front of LCD hinge and in the Specifications Damage due to an incompatible power adaptor is not covered by warranty i CAUTION Never disassemble or service the instrument yourself Do not remove any covers or parts that require the use of a tool to obtain access to moving parts Operators must be trained before being allowed to perform the hazardous operation Unauthorized repairs may damage the instrument or alter its functionality which may void your warranty Contact your local EVOS distributor to arrange for service Q IMPORTANT If you have any doubt about the compatibility of decontamination or cleaning agents with parts of the equipment or with material contained in it contact Technical Support page 71 or your local EVOS distributor for information Sterilization Procedures Q IMPORTANT The EVOS Onstage Incubator should not be subjected to UV sterilization UV degrades many materials including plastic Damage from UV exposure is not covered under the manufacturer s warranty In case hazardous material is spilt onto or into the components of the EVOS Onstage Incubator follow the sterilization procedure as descr
75. ultiple images to build a tiled and stitched image See page 23 for more information e Time Lapse Creates and runs time lapse movies based on user specifications See page 28 for more information 2 Basic Operation Quick Image Capturing Images using Quick Image The EVOS FL Auto Imaging System is a fully automated imaging system controlled by the integrated EVOS FL Auto software The software accessed by the touch screen monitor or the mouse controls the automated X Y axis stage and the objective turret and features imaging and analysis tools Note Refer to the Capture Tab Controls page 10 for more details about the basic image capture functions of the instrument Place the sample on the stage using the appropriate vessel holder For a list of the vessel holders and stage plates available for use with the EVOS FL Auto Imaging System refer to www lifetechnologies com evosflauto or contact Technical Support page 71 1 Onthe Home page click Quick Image Quick Image Options screen opens In the Which Vessel field select the vessel type you are using Using the Where to Start field select the location on the sample vessel you wish to view After selecting the desired area you can choose to see the live image immediately by clicking Live Image Now or complete the remainder of steps and launch the Quick Image Wizard by clicking Launch Wizard If you click Live Image Now you will
76. with the cap provided Dispose of the contents of the waste tray and waste bottle in accordance with good laboratory practices and local state provincial or national environmental and health regulations If potentially hazardous waste is generated when you operate the instrument you must Characterize by analysis if necessary the waste generated by the particular applications reagents and substrates used in your laboratory Ensure the health and safety of all personnel in your laboratory Ensure that the instrument waste is stored transferred transported and disposed of according to all local state provincial and or national regulations IMPORTANT Radioactive or biohazardous materials may require special handling and disposal limitations may apply 67 Electrical Safety Fuses Power Overvoltage Rating 68 AN DANGER ELECTRICAL SHOCK HAZARD Severe electrical shock can result from operating the EVOS FL Auto Imaging System without its instrument panels in place Do not remove instrument panels High voltage contacts are exposed when instrument panels are removed from the instrument WARNING FIRE HAZARD For continued protection against the risk of fire replace fuses only with fuses of the type and rating specified for the instrument DANGER ELECTRICAL HAZARD Grounding circuit continuity is vital for the safe operation of equipment Never operate equipment with the grounding con

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