Tali® Image-Based Cytometer
Contents
1. Increased sensitivity increases recognition of running a sample To adjust the Counting Settings fainter objects in bright field 1 Press the Settings tab on the touch screen operant 2 The Settings screen is displayed Move Sensitivity slider to adjust the sensitivity setting to the desired level Higher circularity requirement recognizes 3 Move the Circularity slider to adjust the more circular objects as cells circularity setting to the desired level 4 Pressing the Default button will return both Default settings to their default values w gt Note In addition to sensitivity and circularity you can also adjust the parameters for the cell size and green and or red fluorescence which are specific to the assay in use For more information see Set gates and thresholds page 22 11 Guidelines for Performing Tali Assays General guidelines Accuracy vs assay 12 speed To obtain the best results follow these recommendations Wear gloves during sample handling Do not touch the optical surfaces of the Tali Cellular Analysis Slides Hold the slides by the edges See page 15 for guidelines on loading the Tali Cellular Analysis Slides Use the Tali Image Based Cytometer at room temperature only 10 35 C The Tali Image Based Cytometer contains delicate optics Place the instrument on a flat dry surface that is free from excessive vibration Do not spill liqui2. 17 Measure Background optional 18 Measure the background optional Measuring the background fluorescence using a non fluorescent sample unstained or non expressing allows you to more accurately set a threshold to distinguish between non fluorescent and fluorescent cells Note that the Background feature is not available for the Quick count assay because this assay does not use fluorescent reagents To measure the background for Tali Viability Apoptosis Green Red and Cell Cycle assays follow the instructions below 1 Press the Background tab The Background screen opens Data Settings A Green Red Data saved as LIVE DEAD 03 Nov 2012 09 40 PM of images to capture 9 fields Press to insert new unstained cell control ss LIVE DEAD_00 amo Conc cells cells Green fluorescent cells 0 00X10e4 cells mL 0 Red fluorescent cells 0 00X10e4 cells mL 0 Total cells 4 86X10e5 cells mL Cell size Layers Background If there are no background measurements for the assay saved on the instrument the In use dropdown menu will display No Background If you choose not to run a background control the instrument will assign a fluorescence threshold for you which you can change manually after performing your assay In use No Background If you have already measured the background the dropdown menu will display the latest background measurement on file TH lice LIVE DEAD_00 me 2 To use a pr
3. recognizes the USB drive a red dot indicates that the USB drive is not inserted into the USB port or that the instrument does not recognize the USB drive After the successful update of the firmware touch OK The Tali Image Based Cytometer will automatically restart to complete the installation of the new firmware 30 Adjust the Camera Alignment Introduction The proper alignment of the two cameras inside the Tali Image Based Cytometer is essential for obtaining the most accurate data from the instrument Follow the protocol below to complete the auto alignment of the cameras so that the fluorescent image is properly overlaid with the bright field image After unpacking the Tali Image Based Cytometer perform the alignment procedure using the Tali Alignment Beads before calibrating the instrument or running any samples for analysis Check the alignment of the Tali Image Based Cytometer every 90 days Materials needed Tali Alignment Beads included in Tali Calibration Beads Cat no T10790 supplied with the instrument or available separately see page 40 Align the cameras 1 Load 25 pL of Tali Alignment Beads into the Tali Cellular Analysis Slide as described on page 15 2 Inthe Settings screen touch Align cameras and initiate the alignment procedure following the directions given on the screen 3 When prompted insert the Tali Cellular Analysis Slide containing the Tali Alignment Beads in
4. Green o mm Layers 2 Move the two blue buttons on the slider bar horizontally to set the lower and upper boundaries for cell size The blue vertical bar on the left determines the minimum cell size and the one on the right the maximum cell size The values set are also displayed under the graph 3 Touch Apply to confirm and return to the previous screen Only cells that fall within the set boundaries are included in the calculations and the screen is updated with the new count To return to the previous screen without setting the threshold for cell size touch Close Continued on next page 22 Run Sample continued Set gates and 4 Toset the threshold for the Green fluorescence touch the Green histogram thresholds continued thumbnail The pop up window displays the of cells vs Fluorescence histogram for Green fluorescence The fluorescence threshold and scaling of the histogram can be set in this pop up Home Date Settings A Green Fluorescence Green Red 129 326 03 Nov 2012 09 59 PM of images to capture new sample cells cells s mL 2 22 mL 0 2 mL 52 508 mL 45 443 Fluorescence RFU mL RFU threshold aii 1968 RFU Bright field fluorescence i z Control soa Eeen a A A pl Close Red J fluorescence pply Layers Zoom Background Sample Select Sample fluorescence or Control fluorescence by touching the correspondi
5. 1 05X10e4 cells mL 2 22 Red only 0 10X10e4 cells mL 0 2 Green and Red 2 42X10e5 cells mL 52 No Green no Red 2 11X10e5 cells mL 45 443 Average cell size 15 um of cells counted 975 Total cell conc 4 65X10e5 cells mL Bright field Circles s Hil alle aanas Green Red J Cell size Green Layers Background Sample amp Note The number of layers that are available for viewing and the labels for those layers depend on the Tali Assay performed In the example above the buttons for the fluorescent channels are labeled Green and Red because the Tali Green Red Assay is selected However if the Tali Apoptosis Assay was selected these same buttons would be labeled Annexin V for the green channel and PI for the red channel 3 Touch the appropriate button on the Layers tab to turn on or off the image for that channel in this example Bright field Green and Red When multiple layers are turned on they appear overlaid on the image 4 To identify the cells counted through a of images to capture 9 fields po particular channel press Circles The Tali Image Based Cytometer circles the cells that were analyzed as follows Blue cells counted in bright field channel Green cells counted in green fluorescence channel Red cells counted in red fluorescence channel Yellow cells counted in both green and red channels Black cells excluded from the count Continued on n
6. CAUTION All biological samples and materials that come into contact with them have the potential to transmit infectious diseases and are considered biohazardous Follow all applicable local state provincial and or national regulations Wear appropriate protective eyewear clothing and gloves IMPORTANT Using a cleaning or decontaminating method other than that specified by the manufacturer may result in damage to the instrument Wipe the touch screen of the Tali Image Based Cytometer using a soft lint free cloth moistened with an LCD cleaning solution Do not apply excessive force during cleaning Wipe the touch screen dry immediately after cleaning e Ensure that the cleaning solution does not enter the power button the power inlet the slide port or the USB ports e Never pour or spray any liquids directly on the instrument to avoid electrical shock when the instrument is plugged in e Do not use abrasive cleaning solutions or material to prevent the touch screen from getting scratched Wipe the instrument case of the Tali Image Based Cytometer using a soft lint free cloth moistened with distilled water Wipe the instrument dry immediately after cleaning e Ensure that water or other cleaning solutions do not enter the power button the power inlet the slide port or the USB ports e Never pour or spray any liquids directly on the instrument to avoid electrical shock when the instrument is plugged in
7. Name later Cell Cycle Green Red Continued on next page 13 Select a Tali Assay continued Selecta Tali 2 To name the sample before performing the assay press Name now Assay continued The alpha numeric keyboard pop up screen is displayed 3 Using the keyboard type the name of the sample series using up to 40 alpha numeric characters and then press Save Each sample run in the series will be appended with a number to reflect the order in which it was run To return to the previous screen without assigning a name to the sample series press Close amp Note If you select Name later the instrument automatically assigns a name for each sample series by date and time You can later rename the individual samples from the Data screen 14 Load Sample Load the Tali The Tali Cellular Analysis Slides are plastic disposable slides that hold the sample Cellular Analysis in two separate enclosed chambers A and B The dual chambers of the slide allow the analysis two different samples or replicates of the same sample Each chamber has a 25 uL sample capacity Follow the guidelines below to load your samples on the chamber slides Slides Do not touch the optical surfaces of the Tali Cellular Analysis Slides Hold the slides by the edges Use 25 uL of sample volume per slide chamber Do not overfill or underfill the slide chambers Pipet the sample at an angle of approximately 80 int
8. Touch screen display Image adjustment focus knob Continued on next page Tali Image Based Cytometer Exterior Components continued Rear and side view The rear and side view of the Tali Image Based Cytometer illustrating the various of Tali Im age parts of the instrument is shown below Based Cytometer Power inlet The power inlet connects the instrument to an electrical outlet through the supplied power cord and the appropriate plug based on the electrical outlet configuration in your country On Off switch The on off switch is located above the power inlet and is as the main power switch It is not necessary to use the on off switch for day to day operation of the instrument On Off switch Image adjustment focus knob Power inlet Tali Image Based Cytometer User Interface User interface The touch screen user interface of the Tali Image Based Cytometer is used to operate the instrument and consists of e The touch screen buttons to operate the instrument e The digital display shows the image of cells and sample data The example below shows the Measure screen of the Green Red assay Green Red of images to capture Tali Cellular Analysis Slides 9 fields Data Data saved as LIVE DEAD Settings vy 03 Nov 2012 09 56 PM Press to insert new sample Green only Red only Green and Red No Green no Red Average cell size of cells counted Tot
9. comma separated value jpg fcs and pdf files for archiving and sample comparisons Tali Image Based Cytometer Exterior Components Front view of Tali The front view showing various parts of the Tali Image Based Cytometer is shown Image Based below Cytometer Power button The power button is used to turn the instrument on and off The blue status light indicates that the instrument is on Touch screen display The touch screen display located in the front of the instrument contains buttons for all the functions needed and displays data from the cell count Slide port The slide port is used to insert the Tali Cellular Analysis Slide containing the sample into the counter for analysis USB port The USB port allows you to transfer and save the cell count data and image to your computer for record keeping and printing purposes The USB drive supplied with the instrument or any other standard USB drive is inserted into the USB port for data transfer See page 28 for instructions on exporting data files Image adjustment focus knob The image adjustment focus knob is used to adjust the image quality to obtain better contrast between the cells and the background as well as between cells that have taken up Tali Image Based Cytometer reagents and unstained cells This is important to obtain accurate cell counts and fluorescence measurements for various Tali Image Based Cytometer assays Power button USB port Slide port
10. precautions Review and follow the safety instructions below Do not install the instrument in heavy humidity such as a greenhouse or an incubator to avoid a danger of electric shock If water or other material enters the instrument the adaptor or power inlet disconnect the power cord and contact a service person For operating environment refer to Appendix A Product Specifications page 38 Do not touch the main plug or power cord with wet hands Always ensure that the power supply input voltage matches the voltage available in your location This instrument is air cooled so its surfaces may become hot during operation When installing the instrument leave a space of more than 10 cm 4 inches around it and do not place any objects between the instrument and the wall Do not install the instrument on a slant or a place prone to vibrations which induces the risk of instrument malfunction or damage of the instrument Never insert any objects into the air vents of the instrument as this could result in electrical shock personal injury and equipment damage Plug the power cord firmly into the wall outlet and AC adapter To avoid potential shock hazard make sure that the power cord is properly grounded Be sure to position the equipment such that it is easy to disconnect the instrument Turn off the instrument before unplugging the power cord and or moving the instrument If the instrument is broken or dropped d
11. 09 56 PM of images to capture 9 fields Press to insert new Sample Conc cells cells Green only 1 05X10e4 cells mL 2 22 Red only 0 10X10e4 cells mL 0 2 Green and Red 2 42X10e5 cells mL 52 508 No Green no Red 2 11X10e5 cells mL 45 443 Average cell size 15 um of cells counted 975 Total cell conc 4 65X10e5 cells mL ale Cell size Red Background 2 To zoom into and out of the selected image move the red dot in the zoom slider bar to the desired magnification 3 To review a different section of an image viewed at 4x or 16x magnification touch the image to display the navigation tool 4 Touch the appropriate area on navigation tool to display the desired section of the image 5 Touch the image outside the navigation tool to hide the tool and review the image of images to capture 9 fields of images to capture 9 fields a V Note The navigation tool is only available for reviewing the images at 4x and 16x magnification Continued on next page 20 Run Sample continued Review layers 1 To review images captured through different channels bright field green fluorescence red fluorescence select the image by pressing the Thumbnail in the Zoom tab and then press the Layers tab Home Data Settings vy Green Red Data saved as LIVE DEAD 03 Nov 2012 09 57 PM of images to capture 9 fields B Press to insert new Sample Conc cells cells Green only
12. 13 Date Align cameras Last calibration 13 Mar 2013 Calibrate Green Red Firmware version Ver 2 1 Update firmware The Settings screen allows you to 16 Jun 2013 07 09 AM Counting settings Sensitivity Increased sensitivity increases recognition of fainter objects in bright field Circularity Higher circularity requirement recognizes more circular objects as cells Default e Calibrate and align the instrument to ensure optimal performance page 31 e Update to install new firmware versions as they become available page 30 e Setup date and time see below e Adjust counting settings sensitivity and circularity page 11 Set up date and The date and time is already preset when you receive the instrument To reset the time date and time use the Date and Time roller wheels that respond to the movement of a finger across the screen as if they were wheels 1 Press the Settings tab on the touch screen to display the Settings screen 2 Select the date and time by bringing the desired value to the center position on the roller wheel located under Screen settings The updated date time is displayed on the top right corner of the screen Once the date time is set there is no need to set it each time the instrument is turned on Screen settings Date 10 Continued on next page Getting Started continued Adjust counting The Tali Image Based Cytometer comes with pre set par
13. Cat no A10798 Viability determines the number and proportion of viable and dead cells using the Tali Viability Kit Dead Cell Red Cat no A10786 which stains the dead cells red Apoptosis distinguishes between apoptotic dead and live cell populations using the Tali Apoptosis Kit Cat no A10788 counts the cells in each population and calculates relative amount of each population in the sample Green Red counts and calculates the population distribution of green and or red fluorescent cells These cells may be expressing GFP or RFP or could be stained with any green or red fluorescent stain The Green Red assay menu includes options for Green alone Red alone and Green Red assays To measure viability in cells expressing fluorescent proteins use the Green Red assay and the Tali Viability Kit Dead Cell Red Cat no A10786 with cells expressing green fluorescent proteins or use the Tali Viability Kit Dead Cell Green Cat no A10787 with cells expressing red fluorescent proteins Quick count provides quick and accurate cell counts without the need for staining your cells and determines the concentration of your sample and the average cell size Note For more information on Tali Assays and assay specific protocols refer to the product information sheets PIS supplied the individual assay kits The PISs can also be downloaded at www lifetechnologies com tali 39 Appendix C
14. Continued on next page Clean the Tali Image Based Cytometer continued Decontaminate the instrument Wipe the instrument case of the Tali Image Based Cytometer using a soft lint free cloth moistened with 70 alcohol Wipe the instrument dry immediately after cleaning Avoid using a bleach solution because it may leave a residue of bleach crystals on the instrument Avoid cleaning the touch screen Ensure that water or other cleaning solutions do not enter the power button the power inlet the slide port or the USB ports Never pour or spray any liquids directly on the instrument to avoid electrical shock when the instrument is plugged in 37 Appendix A Product Specifications Technical Specifications 38 Physical characteristics Technical specifications Optics Tali Cellular Analysis Slide Instrument type Instrument dimensions Weight Operating power Frequency Electrical input Installation site Operating temperature Operating humidity Processing time Sample concentration range Particle cell diameter range Required sample volume Firmware USB Drive Optics Excitation Filters Camera Material Dimensions Chamber volume Benchtop cell counter and suspension cell based assay platform 11 W x 17 D x 11 H 19 4 Ibs 100 240 VAC 2 5 A 120 V 50 60 Hz 12 VDC 13 A Indoor use only Class A Environments i e non resid
15. J 300s 1150 RFU 41501 468d a BALE ogencaini 29 Maintenance Update the Tali Image Based Cytometer Firmware Introduction The update feature allows you to update the Tali Image Based Cytometer when new firmware versions are available This ensures the optimal performance of the instrument and adds new assays and features if available Update the To update the Tali Image Based Cytometer with the latest firmware follow these firmware Steps 1 2 4 Download the latest firmware from www lifetechnologies com tali Extract the files from the zip folder and transfer all files to the root directory of your USB drive IMPORTANT The files must reside individually on the USB drive Do not transfer as a folder or put the files in a folder Touch Update firmware on the Settings screen The Update Dialog Screen is displayed Update firmware Measure Firmware settings EEE Update Last alignment 23 Aug To update firmware insert USB Align camera drive with latest firmware files All awe saved data from instrument will itivity increases recognition of Last calibration 1 JAN 7 remain in memory in bright field Calibrate Green nn Cancel Firmware version Ver 2 1 ee eee bjects as cells Default Update firmware a A Insert the USB drive into the USB port of the Tali Image Based Cytometer and touch Update amp Note A green dot on the Update button indicates that the instrument
16. OEE 41 DOCUMENTALION ANG SUP DON ession a a sien tee aai 43 Oat IS SU POON Ess eaters E AAO A EEE A EE 43 Preface About This Guide Audience and purpose of this manual User attention words Safety alert words The Tali Image Based Cytometer User Guide is for laboratory staff operating maintaining and analyzing data using the Tali Image Based Cytometer The user guide is designed to help you learn how to use the instrument and it includes step by step instructions for instrument set up data collection data handling and instrument maintenance it does not contain assay specific protocols For information on specific Tali Assays and detailed protocols refer to the product information sheets PIS supplied the individual assay kits For your convenience these PISs are included in the Tali Image Based Cytometer USB Drive they can also be downloaded at www lifetechnologies com tali 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 IMPORTANT Provides information that is necessary for proper instrument operation accurate installation or safe use of a chemical Three safety alert words appear in Life Technologies user documentation at points in the document where you need to be aware of relev
17. Ordering Information Accessory Products Tali Image Based Cytometer and related products Tali Image Based 40 Cytometer Assays Some of the components of the Tali Image Based Cytometer are also available separately from Life Technologies These products are listed below For more information see www lifetechnologies com or contact Technical Support see page 43 Product Amount Cat no Tali Image Based Cytometer power cords 1 each T10793 pack of 4 for U S E U U K Australia Tali Image Based Cytometer USB Drive 1 each T10792 Tali Calibration Beads includes 1 tube each of 1 kit T10790 Tali Green Calibration Beads Tali Red Calibration Beads and Tali Alignment Beads Tali Cellular Analysis Slides 50 slides T10794 500 slides T10795 The assay kits listed below have been designed and optimized for use with the Tali Image Based Cytometer For more information see www lifetechnologies com or contact Technical Support see page 43 Product Amount Cat no Tali Viability Kit Dead Cell Red for use with 100 assays A10786 Tali Assays Viability amp GFP Viability Tali Viability Kit Dead Cell Green for use with 100 assays A10787 Tali Assays RFP Viability Tali Apoptosis Kit Annexin V Alexa Fluor 488 100 assays A10788 and Propidum Iodide Tali Cell Cycle Kit 50 assays A10798 Appendix D Safety Safety Information Safety
18. Tali Image Based Cytometer Quick Reference Card QRC 1 each Certificate of Analysis CoA 1 each Tali Calibration Beads 1 kit Tali Calibration Beads are shipped separately and include 1 tube each of Tali Green Calibration Beads Tali Red Calibration Beads and Tali Alignment Beads Examine the instrument carefully for damage incurred during transit Ensure that all parts of the instrument including accessories listed above are included with the product Damage claims must be filed with the carrier the warranty does not cover in transit damage See page 9 for instructions on installing the Tali Image Based Cytometer Visit www lifetechnologies com tali to register your instrument You will be asked to supply the serial number your name and your contact details Registering your instrument ensures that you will receive notifications of software upgrades and information on new assays for use with the Tali Image Based Cytometer Description of Tali Image Based Cytometer Tali Image Based The Tali Image Based Cytometer is a 3 channel bright field green fluorescence Cytometer red fluorescence benchtop assay platform that uses state of the art optics and image analysis to perform assays for cells in suspension including GFP and RFP expression apoptosis cell viability live dead and total cells cell cycle and cell counting assays It is compatible with a wide variety of eukaryotic cells Usi
19. USER GUIDE Tali Image Based Cytometer Catalog Number 10796 Publication Number MANO003766 Revision 2 0 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 control and commercial services such as reporting the results of p
20. aeien acetate 12 Selecta Tall AGS ay sais dime oeesctaincneonecsaindimmnncelaiaceneaie nian ian oneal mon aeons 13 TSO AG DAI PC vars en a S E A A A ONON 15 FOcUS HDI Ere a A a O 17 Measure Backer ound 0P RONAL Asepsis toenulamegeasssbonadenarsoeeantaareusnargaues 18 RUN ch FOES cs ce a nga tan toate Daas ncaa ss A T EE O E 19 Analyze Aes ageing ae Ape Fo ad ees asta 24 MAIN LEN AIMG Cs sacsierchancbacchausiareb suciausharcbaushaunhanebaceinausharabaushaushamebacchausharabaushaunhaebaasiarchares 30 Update the Tali Image Based Cytometer Firmware c ccccsssssesssssseseseseseseseseseseseseeeseeeeeeseeeeeseeeaeaeeeeeees 30 Aust tie Camera A We Mien aiea a E sa roaaneds tyaarkagaieeua may ots a 31 Calibrate the Tali Image Based Cytometer sscsssssssseseseseseseseseseseecseeeseceseseaeseseaeneseeeeeeeeeerseaeaeaeaeeeess 33 Clean the Tali Image Based Cytome te ernst tatcaasncnaey rm cchose a aac tae edad eee 36 Appendix A PROUUCESDECIIICALIONS sasaa eee bew esheets 38 T CCMMUICALS PCCIEICA MOINS suais a a teat aa eaniilcnn debauend A E 38 Append Be Bs ASSN eet en ee Ce ene eR et ne 39 CTC Ott A oe rae re ee hoe A eee eet E E 39 Appendix C Ordering INIOFMAUOMN cect etter etter eieeh eiteniaideeewieeh aioe 40 PRC CESSOUY TE OCICS cata Seco ie saic ch asus A OTE ous las eal cae aotea ind Darina E 40 PAD SVC Ds Sl CU scarcer etc rsa pert eos eee se ees ene ee ee eee 41 DALEY MOL MAO iaia nuaiyeaiasined see E E E iae down ape ene E saat aun
21. al cell conc Cell size Background Conc cells cells 1 05X10e4 cells mL 2 22 0 10X10e4 cells mL 0 2 2 42X10e5 cells mL 52 508 2 11X10e5 cells mL 45 443 15 um 975 4 65X10e5 cells mL Tali Cellular The Tali Cellular Analysis Slides are disposable slides composed of polylactic acid Analysis Slides PLA a low fluorescence plastic Each slide holds the sample in two separate enclosed chambers A and B allowing the analysis of two different samples or replicates of the same sample Each chamber has a 25 uL sample capacity The Tali Image Based Cytometer captures a series of images i e fields of view of the sample in the chamber and then analyzes them using algorithms specific for the Tali Assay selected See page 15 for guidelines on loading the Tali Cellular Analysis Slides Using the Tali Image Based Cytometer Workflow Operation The Tali Image Based Cytometer simultaneously captures a series of bright field principles and fluorescent images i e fields of view of the sample in the Tali Cellular Analysis Slide and uses sophisticated digital image analysis algorithms to determine total and fluorescent cell counts and calculate their concentrations All Tali Assays page 39 share the same basic workflow and user interface but differ in which channels the algorithm is used for analysis Using the touch screen user interface of the Tali Image Bas
22. ameters to match the settings majority of cultured cell types that can be accurately counted without changing any of the default parameters Counting settings of the Tali Image Based Cytometer allows you to adjust the global image analysis parameters for specific or mixed cell types you must be determine these settings empirically From the Settings screen you can adjust the following global parameters for counting e Sensitivity refers to the contrast of the objects from the background Increasing the sensitivity makes the instrument more sensitive to objects in the bright field while decreasing the sensitivity makes the instrument less sensitive which is useful if there is a significant amount of debris in the sample e Circularity is used to determine which objects to include in the measurement based on their roundness Increasing the circularity value requires objects to be rounder for inclusion in the measurement and may be useful if you need to distinguish perfectly round cells from more oddly shaped cells Decreasing the circularity allows objects to be less round for inclusion in the measurement and may be useful if the cell type is not particularly circular or perhaps oddly shaped The default settings for sensitivity and circularity Counting settings are 6 and 8 respectively These values are Sensitivity displayed in red and highlighted in grey in the Counting settings box Counting settings must be adjusted prior to eae
23. ant hazards Each alert word CAUTION WARNING DANGER implies a particular level of observation or action as defined below 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 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 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 Symbols on page 42 Product Information Product Contents Tali Image Based The Tali Image Based Cytometer Cat no T10796 is shipped with the components Cytometer listed below Upon receiving the instrument Registering your instrument Component Quantity Tali Image Based Cytometer 1 each Tali Cellular Analysis Slides box of 50 1 box Tali Image Based Cytometer power cords pack of 4 cords 1 each for U S E U U K Australia Tali Image Based Cytometer USB Drive 1 each includes the user manual
24. ard AS NZS 2064 Limits and Methods Measurement of Electromagnetic Disturbance Characteristics of Industrial Scientific and Medical ISM Radio frequency Equipment This product conforms to UL61010 1 CSA C22 2 No 61010 1 Safety Requirements for Electrical Equipment for Measurement Control and Laboratory Use Part I General Requirements Instruments bearing the TUV symbol are certified by TUV Product Services to be in conformance with the applicable safety standards for the US and Canada This equipment has been tested and found to comply with the limits for a Class A digital device pursuant to Part 15 of the FCC Rules These limits are designed to provide reasonable protection against harmful interference in a residential installation The equipment generates uses and can radiate radio frequency energy and if not installed and used in accordance with the instructions may cause harmful interference to radio communications However there is no guarantee that interference will not occur in a particular installation If this equipment does cause harmful interference to radio or television reception which can be determined by turning the equipment off and on the user is encouraged to correct the interference by one or more of the following measures e Reorient or relocate the receiving antenna e Increase the separation between the equipment and receiver e Connect the equipment into an outlet on a circuit different from tha
25. d Calibrate the green 4 When prompted focus your sample containing the Tali Green Calibration and red channels Beads using the focus knob continued Home Measure Data Green Red 12 10 03 PM Firmware 2 8 iiss Last alignr Focus your sample A using the focus knob 8 10 ognition of Last calibr Calil ii 2 i l i i z Ka Firmware Cancel tognizes Up Default gt Note When correctly focused the Tali Calibration Beads will appear round rather than oblong Correct focus Incorrect focus a as 5 After the beads have been properly focused touch Run The Tali Image Based Cytometer will perform the auto calibration of the green channel 6 When the green channel calibration is complete the cellular analysis slide will be ejected automatically and the red channel calibration dialog box will be displayed Measure Green Red 012 10 09 PM Firmware Insert sample Load red calibration beads Firmware Cancel Continued on next page 34 Calibrate the Tali Image Based Cytometer continued Calibrate the green 7 and red channels continued 10 11 Load 25 uL of Tali Red Calibration Beads into the Tali Cellular Analysis Slide as described on page 15 Make sure that the beads are mixed thoroughly before pipetting Insert the slide containing the red calibration beads into the slide port of the instrument and press Insert sample After
26. djusting the counting settings to optimize the analysis of different sample types Install the Tali 1 After unpacking the instrument place the instrument on a flat level dry surface Image Based 2 Plug one end of the power cord appropriate for your region into the Tali Cytometer Image Based Cytometer 3 Plug the power cord into the electrical outlet Be sure to use only the power cord supplied with your instrument Powering the instrument with an unapproved power cord may damage the instrument 4 Turn on the main power switch i e On Off switch located above the power inlet port See image on page 6 5 When you are ready to use turn on the Tali Image Based Cytometer by pressing the Power button on the front of the instrument see image on page 5 Home screen When the Tali Image Based Cytometer is turned on the instrument initializes and displays the Home screen From the Home screen you can proceed immediately to Tali Assays Quick count Cell Health Cell Cycle or Green Red or use the navigation bars to access saved data Data screen or adjust instrument settings Settings screen Measure Settings vy 09 May 2013 11 36 PM Quick count Cell Health Cell Cycle Green Red Continued on next page Getting Started continued Settings screen Press the Settings tab on the navigation bar to display the Settings screen Measure Firmware settings Screen settings Last alignment 13 Mar 20
27. ds on the surface of the instrument or introduce liquids into its interior The Tali Image Based Cytometer can accurately count cells 5 um to 60 um in diameter The recommended sample concentration range for the Tali Image Based Cytometer is 1 x 10 to 1 x 10 cells mL however the sample concentration does not need to be exact to perform an assay For accurate viability count results ensure the counting area is covered with cell suspension and count cells immediately after staining per the assay protocol Re calibrate the Tali Image Based Cytometer after updating the firmware or when using assay reagents not formulated for the instrument To recalibrate the Tali Image Based Cytometer see page 32 The internal working memory of the Tali Image Based Cytometer is 145 Gigabytes sufficient for storing comprehensive data from 1000 sample runs However we recommend that you save your data to the USB drive after completing each experiment Using the USB drive you may then transfer the data to your PC as described in Export Data page 25 After using Tali Cellular Analysis Slides appropriately dispose of them as biohazardous waste Do not reuse the Tali Cellular Analysis Slides Turn off the Tali Image Based Cytometer at the end of the day The Tali Image Based Cytometer performs counts by simultaneously capturing bright field and fluorescent images of the sample in the chamber slide and then analyzing the cap
28. e list to access hidden data files 3 Select the data file to review by touching the corresponding line on the file list Touch Select Multiple to select more than one file at a time Touch Deselect Multiple to deselect all files Touch Select All to select all files in the list To deselect all files touch any individual file in the list Continued on next page Analyze Data continued Rename data files 1 2 3 Select the data file to rename by touching the corresponding line of the file list The selected line will be highlighted To rename the selected file s press Rename The alpha numeric keyboard screen is displayed Settings A 012 01 07 AM Images jpg Report pdf All formats Layers Zoom Analysis Rename Select all Delete Enter the file name containing up to 40 alpha numeric characters using the touch screen keyboard and then touch Save Touch Close to return to the previous screen without renaming the file Delete data files 1 2 Select the data file s to delete by touching the corresponding line of the file list The files lines will be highlighted Touch Select Multiple to select more than one file at a time Touch Deselect Multiple to deselect all files Touch Select All to select all files in the list To deselect all files touch any individual file in the list To delete the selected file s touch Delete Continued on next page 25 Analyze Data continu
29. ed Review stored data 1 To review the data from counts previously saved in the Tali Image Based Cytometer select the desired file from the Data screen 2 Press Layers Zoom or Analysis tabs to display the corresponding screens Edit notes Edit notes Edit notes Conc cells cells Green only 1 56X10e5 cells mL 73 328 Red only 0 00X10e4 cells mL 0 0 Green and Red 0 10X10e4 cells mL 0 2 No Green no Red 5 72X10e4 cells mL 27 120 Average cell size 15 pm of cells counted 450 Total cell conc 2 14X10e5 cells mL Layers Zoom Analysis L U Cell Size Green Red Analysis Analysis Annotate data 1 To attach a note to your data file select the file to annotate and then touch the Notes box above the Data window to display the alpha numeric keyboard screen Measure Settings vw 03 Nov 2012 01 06 AM Select file s Experiment name 2012 11 01 01 22_1 Jurkat Cell Cycle3 Jurkat Cell Cycle_2 Images EA Jurkat Cell Cycle_i 2012 09 26 23 44_2 2012 09 26 23 44_1 2012_0920_cell cycle_assay vol_7 2012_0920_cell cycle_assay vol_6 Layers Zoom Analysis Rename Select all Delete 2 Enter your comments using the touch screen keyboard and then touch Save Your comments in the Notes box is be attached to the file and shown on all Data tabs i e Layers Zoom and Analysis To return to the previous screen without renaming the file touch Close Contin
30. ed Cytometer you can select the number of images i e fields of view to capture for each experiment and gate the counts by cell size and or fluorescence intensity where applicable After capturing you can select each field of view for review zoom in and out of the selected field of view display channel specific layers i e images captured through bright field green fluorescent and red fluorescent channels or any combination of them and identify the cells counted through each channel Workflow The table below describes the major steps for using the Tali Image Based Cytometer Step Action Page 1 Optional Adjust global counting settings 11 2 Select the Tali Assay you wish to perform 13 3 Load your sample 15 4 Focus the field of view 17 5 Optional Perform a new background measurement or select a 18 previously recorded background not applicable to the Quick Count Assay 6 Capture images of the sample 19 7 Review captured images 20 8 Set gates and thresholds 22 9 Annotate and export your data 24 Note Before using the Tali Image Based Cytometer for the first time you need to calibrate the instrument with the beads provided and align the cameras See page 31 for more information Getting Started Introduction This section provides information on installing your Tali Image Based Cytometer setting the date and time and updating the firmware It also provides information on a
31. ential or light industrial Pollution degree 2 10 40 C lt 80 non condensing 10 seconds to 2 minutes depending on the number of fields captured 1 x 10 1 x 10 cells mL 5 60 um 25 pL Tali Image Based Cytometer Firmware visit www lifetechnologies com tali for updates 4 Gigabyte 3 channels bright field green fluorescence red fluorescence Green channel LED 458 20 nm Red channel LED 530 20 nm Green channel 466 40 EX 495 LP Di 525 50 EM Red channel 543 22 EX 580 LP Di 585 LP EM 1 3 Mega pixels 4x objective 4x or 16x digital Zoom Polylactic acid PLA 110 mm W x 24 mm D x 1 9 mm H 25 uL Appendix B Tali Assays Overview of Tali Assays The Tali Image Based Cytometer incorporates image based cell counting and fluorescence detection algorithms to perform the assays listed below for cells in suspension The instrument captures up to 20 fields of view per sample with each field of view covering 0 233 uL of the sample and presents the relevant data e g cell count vs cell size cell count vs fluorescence in tables and histograms The data from the analysis including the image files can be downloaded to a USB flash drive immediately after the assay and transferred to a computer for sample comparisons Cell Cycle determines the distribution of cells within a population that are in each stage of the cell cycle when used in conjunction with the Tali Cell Cycle Kit
32. escence image for each field of view Images are exported as jpg files Report contains only the aggregate results and calculations from the run such as the total cell count and concentration counts and concentrations of fluorescent cells and their relative abundance in the sample thumbnails of the bright field image of each field of view captured as well as the relevant histograms and instrument settings in a pdf file see page 29 Measure Settings vy 03 Nov 2012 01 06 AM Select file s Select multiple Export Experiment name 2012 11 01 01 22_1 csv fcs Jurkat Cell Cycle3 Jurkat Cell Cycle_2 Images jpg _ Jurkat Cell Cycle_1 Export 2012 09 26 23 44_2 buttons Report 2012 09 26 23 44_1 pdf 2012_0920_cell cycle_assay vol_7 2012 0920 cell cycle_assay vol_6 All formats Layers Zoom Analysis Rename Select all Delete Continued on next page 27 Analyze Data continued Exportdata 1 Insert the Tali Image Based Cytometer USB Drive or any other USB drive into one of the USB ports on the Tali Image Based Cytometer 2 Select the data file to export by touching the corresponding line of the file list The selected line will be highlighted Touch Select all to highlight all files To deselect a file touch the highlighted line on the file list again 3 Touch Data table csv and fcs Image jpg or Report pdf to export the data in the designated formats To ex
33. eviously obtained background measurement select the file name of the measurement from the In use drop down menu To perform a new background measurement load the Tali Cellular Analysis Slide with the unstained cell control insert the slide into the instrument see page 16 and touch Press to insert new unstained cell control To replace a previously obtained background measurement select the file you want to replace from the In use drop down menu insert the Tali Cellular Analysis Slide containing the new unstained cell control into the instrument and touch Press to insert new unstained cell control 3 After the unstained control sample is automatically pulled into the instrument touch Press to run unstained cell control Run Sample Capture images 1 To specify the number of fields of view to capture touch the arrow button on the of images to capture drop down menu and select from the available options of images to capture 4 fields 9 fields 13 fields 18 fields 20 fields J To capture the specified number of fields of view touch Press to run sample While the image capture is in progress an ongoing update is provided by the progress bar Home Green Red of images to capture 9 fields Measuring samples Data Data saved as LIVE DEAD 03 Nov 2012 09 51 PM Settings Press to run sample Green only Red only Green and Red No Green no Red Average cell size o
34. ext page 21 Run Sample continued Set gates and The Tali Image Based Cytometer allows you to set gates or thresholds on cell size thresholds md relative fluorescence from available channels for data collection After capturing images of your sample follow the instructions below to set the thresholds for cell analysis The example below shows how to set gates and thresholds for the Tali Green Red Assay BS Note The procedure for setting gates and thresholds is identical for all Tali Assays but the buttons on the touch screen are labeled according to the specific assay selected For example the histogram thumbnails that are labeled Cell size Green and Red for the Tali Green Red Assay see image below are labeled Cell size Annexin V and PI for the Tali Apoptosis Assay The thin blue lines on the histograms represent the set thresholds a Cell size Green Red and upper boundaries for cell size 1 To set the gating parameters i e lower touch the Cell size histogram thumbnail all The pop up window displays the Cell size vs of cells histogram and the slider bar for setting the lower and upper boundaries for cell size Home D Settings vy Cell size Green Red 22 03 Nov 2012 09 58 PM of images to capture new sample cells cells mL 2 22 mL 0 2 mL 52 508 mL 45 443 of cells Cell size min max mm an Cells counted right fie E
35. f cells counted Total cell conc Cell size Background Conc cells cells 0 0 0 0 0 0 0 0 0 ym 0 0 After capturing the specified number of fields of view the Tali Image Based Cytometer automatically ejects the slide and provides data from the analysis of captured images in the Data window of the Sample tab The data include average cell size number of cells counted total cell concentration as well assay specific data and histogram plots In this example the Tali Green Red Assay specific data include the concentration percentage and number of cells showing green fluorescence only red fluorescence only both green and red fluorescence and no fluorescence as well as the histograms for cell size and green and red fluorescence intensity Green only Red only 4 91X10e4 cells mL Green and Red No Green no Red Average cell size of cells counted Total cell conc Cell size cells 22 103 694 470 cells 2 8 54 36 D Red Conc 1 05X10e4 cells mL 3 31X10e5 cells mL 2 24X10e5 cells mL 14 um 1289 6 14X10e5 cells mL Green To run the next sample insert the new slide and repeat the procedure Continued on next page 19 Run Sample continued Review captured 1 To select a captured field of view for review press the thumbnail of the image images in the Zoom tab Home Data Settings vy Green Red Data saved as LIVE DEAD 03 Nov 2012
36. he Tali Image Based Cytometer counts the cells in the chamber opposite the loading direction For example to count the sample in Chamber A insert the Tali Cellular Analysis Slide into the slide port of the instrument with the Chamber B side first until it stops see figure below Do not forcefully push the slide any further instrument 3 Touch Press to insert new sample the slide will automatically be pulled into the instrument Press to insert new sample gt Note If you are performing a background measurement the button for inserting the sample will read Press to insert unstained control see page 18 16 Focus Image Focus the field of 1 Before running your sample focus your cells using the image adjustment view focus knob on the right side of the instrument 2 Press Zoom and select either 4x or 16x magnification enhancement to review the image e Correctly focused images have uniformly dark colored cells surrounded by bright halos see example on the left below e Cells may be undercounted when the transition between the background and the edges of the cells are fuzzy and the cells have undefined boundaries see example in the middle below e Cells maybe overcounted when they have bright centers and dark perimeters see example on the right below Image is correctly focused Incorrect cells may be undercounted Incorrect cells maybe overcounted Continued on next page
37. isconnect the power cord and contact a service person Do not disassemble the instrument Use only authorized accessories adaptor power cord and USB drive For operating environment see Appendix A Product Specifications page 38 If the instrument emits smoke disconnect the power cord from the wall outlet and contact a service person Continued on next page 41 Safety Information continued 42 Symbols FCC compliance The symbols used on the Tali Image Based Cytometer are explained below Used on the instrument to indicate a warning caution risk of danger Consult the manual to avoid possible personal injury or instrument damage Protective conductor terminal main ground WEEE Waste Electrical and Electronic Equipment symbol indicates that this product should not be disposed of in unsorted municipal waste Follow local municipal waste ordinances for proper disposal provisions to reduce the environmental impact of WEEE Visit www lifetechnologies com weee for collection and recycling options 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 This instrument has been tested to and complies with stand
38. ng button on the bottom left A filled in button indicates the histogram currently being displayed To set the x axis scale press on the minimum or maximum RFU value relative fluorescence unit and type in a new value in the popup key pad To set the threshold move the blue button on the slider bar to the desired fluorescence Alternatively touch the RFU threshold button and type in the threshold value in the popup key pad Touch Apply to confirm this setting and return to the previous screen Only cells above the threshold i e to the right of the blue boundary line are counted as fluorescent and the screen is updated with the statistics To return to the previous screen without setting the threshold touch Close To set the threshold for Red fluorescence touch the Red histogram thumbnail and repeat the above procedure amp Note When using the Tali Cell Cycle Assay the Propidium Iodide PI histogram will have three individual thresholds which are used for partitioning the cell population into the stages of the cell cycle Each threshold can be set as described above and the color of each partition will correspond to the appropriate data set in the data table BS Note Biological molecules found within cells fluoresce upon excitation and result in background fluorescence Because the Tali Image Based Cytometer is a highly sensitive instrument it detects this background fluorescence and displays it as a peak clo
39. ng the only 25 uL of sample volume the Tali Image Based Cytometer takes 10 seconds to 2 minutes for a typical assay depending on complexity of the assay and number of fields captured In addition to the bright field channel the Tali Image Based Cytometer features two fluorescent channels green and red enabling it to simultaneously count green or red fluorescent stains as well as cells expressing GFP and RFP The Tali Image Based Cytometer offers an intuitive user interface and provides the option to save data and generate a report which can then be transferred to a PC using the USB drive supplied with the instrument or available separately The Tali Image Based Cytometer is supplied with disposable Tali Cellular Analysis Slides see page 7 which are also available separately see page 40 for ordering information See Instrument Exterior Components page 5 for details on various parts of the Tali Image Based Cytometer Features Important features of the Tali Image Based Cytometer are e Provides a user friendly benchtop design for simple fast and highly accurate three parameter population analysis e Uses the Tali Assays optimized for the Tali Image Based Cytometer see page 39 e Uses disposable Tali Cellular Analysis Slides that eliminate washing steps and cross contamination between samples see page 7 e Presents comprehensive data with graphic reports and allows the export of data as cSv
40. o the half moon shaped sample loading area see figure below The sample is loaded into the chamber through capillary action Take care to avoid forming bubbles in the sample The Tali Image Based Cytometer counts the cells in the chamber opposite the loading direction For example to count the sample in Chamber A you need to insert the Tali Cellular Analysis Slide into the instrument with the Chamber B side first see figure below until it stops For inserting the Tali Cellular Analysis Slide into the instrument see page 16 IMPORTANT The Tali Cellular Analysis Slides are specifically designed for use with the Tali Image Based Cytometer exclusively Use of other slides result in inaccurate cell counts and can damage the Tali Image Based Cytometer The Tali Cellular Analysis Slides are supplied with the instrument and are also available separately see page 40 for ordering information Note The slide port button located on the top right side of the a touch screen is used only for opening the slide port to eject the Tali Cellular Analysis Slide in case of an error Pressing this button will not insert the slide into the instrument or eject it after a run is completed Continued on next page 15 Load Sample continued Insert the Tali 1 Load 25 pL of your sample per slide chamber into the Tali Cellular Analysis Cellular Analysis Slide as described on page 15 Slide into the 2 T
41. one 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 17 June 2013
42. port data in all these formats touch All formats Settings vy 03 Nov 2012 01 06 AM Select file s Export Experiment name Data table 2012 11 01 01 22_1 csv fcs Jurkat Cell Cycle3 Jurkat Cell Cycle_2 Images Jurkat Cell Cycle_1 Export 2012 09 26 23 44_2 buttons Report 2012 09 26 23 44_1 oo 2012 _0920_cell cycle_assay vol_7 All formats 2012 _0920_cell cycle_assay vol_6 Layers Zoom Analysis Rename Select all Delete a SS Maaalala 4 Transfer the files on the USB drive to your PC You may open the exported files using the appropriate programs 28 Analyze Data continued Cell analysis report The image below shows an example of a data analysis report exported as a pdf Tali image based cytometer Cell analysis report Var 2 1 Foa Nama 2073 04 07 01 12_1 Exzpornmant Data Annn y onia Na ma Sen mitimity Circu larity Ragenn A Ragion B Ragion C Fagien D 2013 04 07 01 15 28 gt Can cycn Ne Background 76 8 Conc 1 1721085 come mL 1 17 10e6 come mL SASS T05 coms ml 6 B6210e5 enms ml Average com smin p F of cala countaa 11706 Tots call conce a T g 7 of calla Fiucraszamna R F uj RFU threshold invitrogen 23551 Willow Creek Acad Eugena Oregon Sra02 752 peat 54i web ead 2 578170e6 come mL em Can sizeijiin Can ning minimas 511 Propidium loaiaa Fi Bore a E RFU RFU threat 520
43. sest to the 0 RFU value To eliminate the background fluorescence from your calculations adjust the threshold to exclude this peak 23 Analyze Data 24 Introduction The Tali Image Based Cytometer working memory holds 145 Gigabytes of data sufficient for storing numeric and graphic data files from 1000 sample runs You can access the stored data files through the Data tab where you can analyze annotate rename or delete them You can also export the data table containing comprehensive information on individual cells as a csv file or as an fcs file for data analysis using flow cytometry software The overlaid image histograms and the data table for a run can also be exported as a single page pdf report and the individual raw images captured during the run as separate jpg files Select data files 1 Touch Data to navigate to the Data screen which displays the stored data files in a list on the right side of the screen The most recent data file is displayed in the first line of the list Measure Settings vy 03 Nov 2012 01 06 AM Select file s Select multiple Export Experiment name Data Se A l table 2012 11 01 01 22_1 Jurkat Cell Cycle3 Jurkat Cell Cycle_2 Jurkat Cell Cycle_1 2012 09 26 23 44_2 2012 09 26 23 44_1 2012_0920 cell cycle_assay vol_7 2012 0920 cell cycle_assay vol_6 Zoom Analysis Rename Select all Delete 2 Drag the red button on the scrollbar vertically to move up and down th
44. t to which the receiver is connected e Consult the dealer or an experienced radio TV technician for help Documentation and Support Obtaining Support Technical Support Safety Data Sheets 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 43 Headquarters 8 5791 Van Allen Way Carlsbad CA 92008 USA Ph
45. the green and red fluorescent channels of the Tali Image Based Cytometer sets the dynamic range of the instrument The calibration is most effective when performed after the alignment sequence Re calibration of the fluorescent channels is recommended at least once per year Materials needed Tali Calibration Beads Cat no T10790 supplied with the instrument or available separately see page 40 gt Note Tali Calibration Beads contain separate vials of green and red beads necessary for calibrating the green and red channels of the Tali Image Based Cytometer Calibrate the green 1 Touch Calibrate GFP RFP on the Settings screen J I and red channels The Calibration Dialog Box opens Calibrate GFP REP Home Measure vy 12 01 14AM Firmware Insert sample Last alignr i I Load green B 9 10 A calibration beads vaa cognition of Last calibre Calil D 9 10 Firmware Cancel ognizes Up Default 2 Load 25 uL of Tali Green Calibration Beads into the Tali Cellular Analysis Slide as described on page 15 Make sure that the beads are mixed thoroughly before pipetting 3 Insert the slide containing the green calibration beads into the slide port of the instrument and press Insert sample After the slide is pulled in you will be given a live fluorescent image of the green calibration beads Continued on next page 33 Calibrate the Tali Image Based Cytometer continue
46. the slide is pulled in you will be given a live fluorescent image of the red calibration beads When prompted focus your sample containing the Tali Red Calibration Beads using the focus knob After the beads have been properly focused touch Run The Tali Image Based Cytometer will perform the auto calibration of the red channel After the calibration process is completed touch OK to return to the Settings screen There is no need to recalibrate each time the instrument is turned on Measure Al Green Red 012 10 12 PM NEIS Last alignr A Finish calibration Last calibre Calil a OK Firmware hs Note For optimal calibration follow the guidelines below e Load slide with calibration beads just at the time of use to prevent drying of solution in slide e Allow the beads to settle in the slide before pressing run There should be no movement of beads on the image e The entire calibration can take up to 20 minutes to complete 35 Clean the Tali Image Based Cytometer 36 Introduction Clean the touch screen Clean the instrument case We recommend that the Tali Image Based Cytometer be cleaned periodically to prevent the buildup of dust and dirt that might reduce its performance and cause contamination CAUTION To avoid electrical shock always turn off the Tali Image Based Cytometer and unplug the power cord before cleaning or decontaminating the instrument
47. to the instrument Measure Green Red Firmware settings Last alignment 13 Mar 2013 Align cameras Last calibration 13 Mar 2013 Calibrate Green Red Firmware version Ver 2 1 Update firmware 012 09 47 PM Firmware Insert sample Load alignment beads cognition of Continued on next page 31 Adjust the Camera Alignment continued Align the cameras 4 When prompted focus your sample containing the Tali Alignment Beads continued using the focus knob Home Measure Settings vy Green Red 012 09 48 PM Firmware Ta Last alignr A Focus your sample using the focus knob ognition of Last calibr Calil H o 10 Firmware OK Cancel ognizes Up Default 5 After the beads have been properly focused touch Run The Tali Image Based Cytometer will perform an auto alignment 6 Touch OK to confirm the correct alignment of the cameras To cancel the alignment touch Cancel Home Measure Settings vy Green Red Firmware Last alignr A Last calibr Calil Firmware Up 012 09 48 PM Rotate Move OK Cancel ognizes ognition of ee Ho 10 Default amp Note You can also manually adjust the cameras by positioning the green circles over the corresponding Tali Alignment Beads using the directional keys on this screen but this is not recommended 32 Calibrate the Tali Image Based Cytometer Introduction Calibrating
48. tured fields of view using assay specific algorithms You can obtain higher accuracy by capturing more images and thus analyzing more cells The theoretical CV coefficient of variation decreases as the number of cells analyzed is increased If very high accuracy is not required you may choose to capture fewer fields of view to obtain faster assay speeds You can calculate the theoretical CV using the following equation nn where n is the number of cells counted Select a Tali Assay Note For an overview of Tali Assays see page 39 For detailed Tali Assay protocols refer to the product information sheets PIS supplied the individual assay kits For your convenience these PISs are included in the Tali Image Based Cytometer USB Drive they can also be downloaded at www lifetechnologies com tali Select a Tali Assay 1 To select a Tali Assay press the appropriate assay button Quick count Cell Health Cell Cycle or Green Red on the Home screen In the example below Cell Health assay is selected which gives the Viability and Apoptosis assay options Home Measure Settings vy 31 Oct 2012 07 31 PM Please choose assay Quick count Viability Apoptosis Cell Cycle Green Red Next Apoptosis assay is selected from the options giving you the option to name the sample now or later Measure Settings vy 31 Oct 2012 07 32 PM Annexin V Name sample series Quick count Name now
49. ued on next page 26 Analyze Data continued Data export options The Tali Image Based Cytometer is designed for stand alone use it does not require the use of an external computer However to archive data and generate reports you may transfer the data stored in the instrument to your computer using the USB drive The Tali Image Based Cytometer allows you to export the Data table csv Flow Cytometry Standard fcs Images jpg and the final Report pdf separately or all at once using the Export buttons see image below The Data table contains comprehensive information about the count in a spreadsheet format as a csv file comma separated value including size and fluorescence intensity of each individual particle cell in the sample You can import the csv file into any spreadsheet program The Flow Cytometry Standard fcs file 3 0 is the standard data format used for analysis in common flow cytometry software This file contains the fluorescence intensity of each individual cell and uses cell size as a surrogate for forward scatter Because the Tali Image Based Cytometer is an imaging system there is no side scatter so this parameter will not be displayed Choosing Images exports a single image in each channel collected for each field of view For example if you choose Images for an Apoptosis assay with 9 fields of view the instrument will export a bright field image a green fluorescence image and a red fluor
50. urchaser 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 2013 Life Technologies Corporation All rights reserved Contents FRET ACE sisusento nausea ta nsisaaieraetasia agate tanas a ts tat heparin temaitet 2 PRDOUE TIS KSI CO ennie N sun tesoodenonals nea saiadesbanteausdss 2 ProdUctCINTOr MAON ii taescarete us cartes Sa eann canes oc eneusans aaa aneeaanctnascaeieme tae 3 Product Comets s c sccscsssemersaqucanssansnnraieaansineieeynaaaeeeeeere R 3 Description of Tali Image Based Cy Om erent octane gs5 sc atacand euch ceric ot eee arte ues a 4 Tali Image Based Cytometer Exterior Components ccscsesssesssseseeeseeeseeeeeseseseseseseeeeeeeseeeseeeseaeeeneneeeee 5 Tali Cellular Analysis SUGGS sena a a E at ae ete ead aaaaemneaadeaa 7 Using the Tali Image Based Cytometer ccccccssccceeeeeeeesseceeeeeeeeeeeeeessseeeeeeeeeeneeeeeeas 8 WV OE OWN airtel waa ta eee steele st eae aa ates ea sta cates ent anauetaae ua aedints 8 CENE tAr TE e E deta E dng anesa edhe T E e E A 9 Guidelines for Performing Tali ASSAYS cciccstachacevnsesossnderay aniatinniectisavasetinsgdlentsaeedsnina acta aunb
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