Summary of Contents for Thermo Scientific EVOS M5000
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EVOS M5000 Imaging System ™ For Fluorescence and Transmitted Light Applications Catalog Number AMF5000 Publication Number MAN0017563 Revision A.0 For Research Use Only. Not for use in diagnostic procedures.
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Information in this document is subject to change without notice. Revision history MAN0017563 Revision Date Description 07 Aug 2018 New user guide for the EVOS M5000 Imaging System ™ Important Licensing Information These products may be covered by one or more Limited Use Label Licenses. By use of these products, you accept the terms and conditions of all applicable Limited Use Label Licenses.
Contents About this guide ........................3 1. Product information ......................4 Product description ............................4 Standard items included ..........................5 Instrument exterior components and mechanical controls ............... 6 Graphical user interface (GUI)........................8 2. Installation ........................9 Operating environment and site requirements ................... 9 Prepare for installation ..........................
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9. Instrument care and maintenance ................... 58 General care ..............................58 Objective lens care ............................58 Stage care ................................ 59 Decontamination procedures ........................59 Change EVOS ™ LED light cubes ......................... 60 Change the objectives ........................... 61 Calibrate the objectives ..........................62 Appendix A: Troubleshooting ....................
About this guide Audience This user guide is for laboratory staff operating, maintaining, and analyzing data using the Invitrogen ™ EVOS ™ M5000 Imaging System. User attention Two user attention words appear in this document. Each word implies a particular level of observation or action as described below.
1. Product information Product description EVOS M5000 The Invitrogen ™ EVOS ™ M5000 Imaging System (Cat. No. AMF5000) is a fully ™ integrated, digital, inverted imaging system for four-color fluorescence and Imaging System transmitted-light applications. It combines precision optics, a five-objective turret, an 18.5 inch high-resolution LCD display (1920 ×...
Standard items included EVOS M5000 • EVOS ™ M5000 Imaging System, includes the following components and pre- ™ installed accessories: Imaging System 18.5 in articulated LCD monitor (1920 ×1080 pixel resolution) Embedded PC (4 GB RAM) Light cube shipping restraint (remove before use) Stage lock pin (remove before use) EVOS Condenser Light Shield, 110 mm...
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Rear view LCD monitor Network port Mechanical X-Y stage USB-A 2.0 ports (4×) Focusing knobs USB 3.0 port (1×) Power switch Stage X-axis positioning knob 4-pin power input port (12 VDC, 5 A) Stage Y-axis positioning knob* Display port (video output) * Not visible in this perspective.
Graphical user interface (GUI) GUI layout The GUI of the system consists of the Viewing area on the left and Capture and Review tabs and the Settings and Virtual keyboard buttons on the right. Each tab and button opens the controls necessary to execute the selected function. Viewing area: Displays the sample.
2. Installation Operating environment and site requirements • The dimensions of the EVOS M5000 Imaging System are 18 × 23 × 18 in ™ (W×H×D) (46 × 59 × 46 cm). The system requires a benchtop of approximately 36 × 36 in (92 ×92 cm). •...
Prepare for installation Receive and inspect Verify that the items shown on the shipping list are the same items that you ordered at the time of purchase. the shipment Carefully inspect the shipping containers and report any damage to the Thermo Fisher Scientific service representative.
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Remove shipping The EVOS ™ M5000 Imaging System is equipped with two shipping restraints (stage lock pin and light cube shipping restraint) to protect the instrument from restraints shock and vibration during transport. You must remove the shipping restraints before you power on the EVOS ™...
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Blank light cube Thread hold Screws Using the light cube tool, loosen the two screws that secure the blank light cube to the instrument. You do not need to remove the screws. Screw the light cube tool to the thread hold on the light cube, then lift the blank light cube up and out of the light cube turret.
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Secure the UV light shield mount to the top of the condenser light shield using the two screws supplied with the UV shield assembly (not the protruding screws on the mount). Clip the condenser light shield with the attached UV light shield mount back onto the condenser head.
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Power ON the Turn the instrument power switch (located on the back; see page 7) to the ON position. EVOS M5000 ™ Imaging System When the Capture tab is displayed, the EVOS ™ M5000 Imaging System is ready to use. IMPORTANT! All shipping restraints must be removed before turning on the EVOS ™...
3. Capture images Overview Capture tab The basic functions of the EVOS ™ M5000 Imaging System, such as viewing the sample, setting optimal focus, and capturing and saving images are performed in the Capture tab, which is the first screen after start-up. Workflow Select objective and light source ...
Capture images Select objective and Place the vessel containing your sample on the stage using the appropriate vessel holder. light source Note: When capturing images in fluorescence channels, place the light shield box on the stage, over the sample. This is important for optimal image quality. Set the magnification using the objective selection wheel (page 6).
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Adjust brightness For Brightness mode, select Simple or Actual. Simple mode allows you to control Brightness as a single parameter. Actual mode allows you to adjust Light (i.e., LED intensity), Exposure, and Gain parameters individually. Adjust brightness using the Brightness controls: •...
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Focus on the Use the Coarse and Fine focus sliders to focus on the sample. sample Alternatively, click Coarse and Fine autofocus to autofocus on the sample. To limit the autofocus algorithm to a specific region along the Z-axis, use the Autofocus Maximum and Autofocus Minimum controls (blue triangles on the Coarse focus slider) to set the upper and lower bounds.
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Find the region of Use the X-axis and Y-axis positioning knobs (page 6) to move the sample stage and bring the region of interest into the field of view. interest To zoom in and out of the Viewing area, use the Zoom slider. The zoom range is 100% to 1000%.
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Capture image in a Ensure that the Channel you want to capture is selected. single channel When you select a Channel, the corresponding Capture channel box is automatically checked. Note: If you have exited the Live mode by clicking the Channel button again, the current channel remains selected, as indicated by the blue line underneath the corresponding channel button.
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To capture the same field of view in another channel, select the desired Channel. If needed, re-adjust the brightness and focus, then click Capture. The viewing area shows the image captured in the new channel superimposed on the image captured in the previous channel. A thumbnail of the image captured in the new channel is displayed along with the thumbnail of the image from the previously captured channel.
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Capture multiple To capture multiple channels simultaneously, select the desired channels by checking corresponding boxes. channels simultaneously If needed, adjust brightness and focus for each of the selected channels as described. Click Capture to acquire an image in each of the selected channels. The Viewing area shows a multicolor overlay of the images captured in each selected channel.
Analyze and annotate captured images Display settings Display settings and analysis tools allow you to change image display settings for live and captured images in the Viewing area, and to analyze and annotate and analysis tools captured images. Hover the pointer over the Viewing area to reveal the buttons for Display Settings and Analysis Tools.
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Display grid Click the Grid button to superimpose a grid over the Viewing area. To change the grid size, click the Grid Settings button (the arrow on the Grid split button) to open the Grid Settings tool. Select the Size for the grid: 200 µm×...
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Display histogram Click the Histogram button to open the Intensity Histogram plot. Intensity Histogram plot shows the Pixel count vs. Intensity data of the image displayed in the Viewing area. Click the Histogram button again to close the Intensity Histogram plot. You can also close to plot by clicking the X on the plot.
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Add and show Click the Measurement and Annotations Tools button (the arrow on the Show Measurements and Annotations split button) to open the measurements and measurement and annotations tools. annotations Using the Draw… tools, draw a rectangle, ellipse, polygon, line, or a free- form shape over the region of interest on the Viewing area.
Show cell count Show Cell Count Show Cell Count function allows you to count objects in captured or saved images. function Hover the pointer over the Viewing area to reveal Display Settings and Analysis Tools, then click the Show Cell Count button to display Auto Count and Manual Count options in the tabs area.
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To identify the target objects to include in your count, click Target, then click and drag to draw a circle (green) around a representative target. Note: For best results, follow these guidelines when identifying target objects: • When selecting objects, circle the entire object and include a slight border around it.
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To count closely grouped objects that are touching or overlapping as distinct objects, select from the Split Cells options: None: Touching or overlapping objects are not counted separately. Shape: Distinct objects are identified based on shape. Intensity: Distinct objects are identified based on intensity. When you are finished defining the target and background areas, the software automatically counts the objects based on your examples.
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10. The Refine section displays a histogram plot showing Count versus Intensity, Area, or Circularity. In this example, Intensity is selected. 11. To refine your count, select Intensity, Area, or Circularity, then move the handles of the gate bars in the desired direction to set the upper or the lower boundary for the selected parameter.
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Perform Manual Click the Show Cell Count button, then select Manual Count. Count Select the Channels to display in the Viewing area for manual count. You can select multiple channels, provided that they contain captured images. In this example, only DAPI and TX Red channels contain captured images, and both are selected for the manual count.
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Click on the Label number to select a label, then left-click at each point on the Viewing area to tag the items in that label category. You can switch labels as desired. As you tag the objects onscreen with the selected label, the system keeps a running tally of the counts with percentages for each label assigned.
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Save count results When finished with your auto or manual count, click Save to open the Save Composite Image dialog, then navigate to the folder where you want to save your count results. Select Save as screenshot to preserve a detailed account of your count results as an image.
4. Save captured images Save Save images After capturing an image in a single channel or a set of images in multiple channels, click Save to open the Save dialog, then navigate to the folder where manually you want to save your captured images. Select Save individual channels to save images captured in different channels individually.
Quick Save images Optional : Enable Quick Save function allows you to set the save options in advance and save your captured images with a single click directly from the Capture tab or automatically Quick Save without having to click Save after each capture. Click the Quick Save Settings button (the arrow on the Save split button) to open the Quick Save Settings dialog.
5. Capture and save time lapse images Time Lapse tool With the Time Lapse tool, you can set up your cells and program the EVOS ™ M5000 Imaging System to capture images (including Z-Stack images) at given intervals over a time period based on your specifications.
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Adjust brightness and focus for each selected channel as described in “Capture images” (page 16), then click Next. Note: When the sample is focused and ready, tighten the stage brakes (page 6) to prevent the stage from drifting during the time lapse routine. If you have opted to run Auto Focus, select the Auto Focus Channel to use.
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If you have opted to capture Z-Stack images, define the In Focus, Top, and Bottom positions for the Z-Stack image set: If not already on, click the Light button to illuminate the sample as you locate the boundaries and the In Focus position of the Z-Stack. b.
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12. Enter the Total Time for Run 1 into the corresponding Hours, Minutes, and Seconds fields. You can create multiple Runs for a Time Lapse Routine, each with its own duration and image capture frequency. 13. Set the Image capture options for Run 1. Available options are: •...
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14. Optional: Click Add Run to add another Run to the Time Lapse routine, then define the Run options as described for Run 1. 15. When finished with adding Runs, click Next to proceed to Save options. 16. Click the Browse button to open Choose Save Folder dialog, go to the destination folder for the Time Lapse images, then click Select Folder.
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18. Click Start to run the Time Lapse routine. The system provides information about the progress of the Time Lapse routine as it captures the images based on the routine specifications. 19. When the Time Lapse routine is completed, the Review tab opens to the files captured in the routine.
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Run a saved time Each time you run a Time Lapse routine, the system saves the specifics of the routine, which you can then recall and run with a new sample. lapse routine On the Capture tab, click Time Lapse to open the Time Lapse tool. Click Load Routine to open the Choose Routine dialog.
6. Capture Z-Stack Z-Stack tool Z-Stack tool allows you to capture multiple images of a selected field at different focal planes along the Z-axis and combine them to generate a final image with a greater depth of field than any of the individual source images. Capture Z-stack images On the Capture tab, click Z Stack to open the Z-Stack tool.
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If not already on, click the Light button to illuminate the sample, then adjust brightness and focus using the corresponding sliders on the Capture tab. The focus position you set in this step is the initial Current Z position. To accept the Current Z as the In Focus position, click Set for In Focus. To change the In Focus position, move the Coarse and Fine focus sliders on the Capture tab to focus on the sample, then click Set.
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Click the Browse button to open Choose Save Folder dialog. Go to the destination folder for the Z Stack images, then click Select Folder. 10. Select the File Type: PNG or TIFF. 11. Select Generate Projection Image to generate a projection image.
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13. When the Time Lapse routine is completed, the Review tab opens to the files captured in the routine. EVOS M5000 User Guide ™...
7. Review saved images Overview Review tab The Review tab allows you to review still images, including those captured in Time Lapse routines and Z-Stacks. You can also use this tool to annotate the saved images, and to re-save or delete saved files. For the descriptions of the controls available in the Review tab, see page 71.
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Review images To display the folder/image preview in list format, click the List view button. To display the folder/image preview in grid format, click the Grid view button. If needed, use the scroll bar to search the preview list for the desired image file. Click the image to select it.
Analyze saved images Analyze and Hover the pointer over the Viewing area to reveal the buttons for Display Settings and Analysis Tools. annotate saved images The analysis and annotations tools function in the same way in the Capture and Review tabs: •...
8. Adjust instrument settings Overview Settings tab The Settings tab allows you to assign objectives to the objective turret, to calibrate objective magnification, to set image options (highlight saturated pixels and adjust white balance), define savings options for TIFF files, add and remove LED light cubes, and to connect to a Wi-Fi network and to map network drives.
Adjust objective settings Assign objectives After adding a new objective to the objective turret or replacing an older objective, assign the new objective to the appropriate turret position. For instructions on how to remove an objective, see “Change the objectives”, page 61. Go to the Settings ...
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Calibrate objective Go to the Settings Objectives, then click the Calibrate icon next to the newly installed objective to launch the Objective Calibration tool. magnification Mount the EVOS ™ Calibration Slide in the vessel holder, select the objective you want to calibrate, then click Calibrate Objective. Follow the onscreen instructions to calibrate the objective magnification for the selected objective.
Calibrate white balance Calibrate white Calibrate White Balance calibrates color channel lighting. balance Go to the Settings Visuals tab. Click Calibrate White Balance to open the White Balance Calibration tool. Select a 20X or higher objective, then set the phase ring to Brightfield using the Phase annuli selector (page 6), Remove any samples from the X-Y stage, then center the stage using the stage positioning knobs (page 6).
Set saturated pixel options Highlight saturated Highlight saturated pixels function displays overexposed pixels on an image with the user-defined color, which provides a visual aid for optimal illumination when pixels adjusting the brightness settings. Go to the Settings Visuals tab, then click the Highlight Saturated Pixels checkbox.
General settings Define saving Go to the Settings General tab. options for TIFF files To save TIFF files in a Microsoft compatible format, check the corresponding box. To save TIFF files uncompressed, check the corresponding box. When finished with your selections, click Done. Reverse focus Go to the Settings ...
Configure network settings You can connect the EVOS M5000 Imaging System to a network via an Ethernet ™ cable or Wi-Fi adaptor and save captured images directly to shared folders on the network. You can also connect to Thermo Fisher Cloud to store your files. Connect to a Wi-Fi Click the Settings button to open the Settings tab.
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Map network drive Go to the Settings Network tab, then click Map Network Drive. Map Network Drive dialog opens. Select the drive you want to map from the Drive menu. Click Browse to open the Browse for Folder dialog. Navigate to the folder you want to map, then OK.
9. Instrument care and maintenance General care • When cleaning optical elements, use only optical-grade materials to avoid scratching soft lens coatings. • Use the appropriate cleaning solutions for each component, as indicated in the Decontamination Procedures below. • If liquid spills on the instrument, turn off the power immediately and wipe dry.
Stage care • Clean the X-Y stage as needed with paper towels or Kimwipes ™ tissues dampened with 70% ethanol. • When moving the EVOS ™ M5000 Imaging System, be sure to lock the X-Y stage using the Shipping Restraint as shown on page 61 to prevent the stage from sliding.
Change EVOS LED light cubes ™ To customize your EVOS M5000 Imaging System, you can add and remove LED ™ light cubes to fit the instrument’s functionality to your own specific research needs. Each LED light cube is coded to allow the EVOS ™...
Change the objectives To customize your EVOS M5000 Imaging System, you can add and remove ™ objectives to fit the instrument’s functionality to your own specific research needs. Procedure for Remove the objective you want to replace from the objective turret. You may objective change need to move the stage so that the objectives are accessible.
Calibrate the objectives Calibrate objective magnification allows the calibration of the field of view, parfocality, and parcentration parameters of the selected objective. When calibrated, parfocality ensures that the sample stays in focus when the objective is changed, and parcentration ensures that an object in the center of the field of view stay in the center of the field no matter which objective is being used.
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Find a calibration circle that fits entirely in the field of view. Focus on the calibration circle using the focus controls. Click and drag the green calibration lines to align them along the outer borders of the calibration circle. When finished, click Save to complete the calibration. Repeat the calibration process for each additional objective to be calibrated.
Appendix A: Troubleshooting Note: For additional technical support, contact your local EVOS ™ distributor. If you do not have your distributor information, visit thermofisher.com/evos or contact Technical Support (page 86). Image quality issues Problem Possible solutions Image is too dim (at higher Remove condenser slider, if one is in place.
Software interface issues Note: We recommend keeping the EVOS M5000 Imaging System up to date with the latest software. ™ Problem Possible solutions Image does not respond to changes in Click the Light button to return to live observation. focus or stage position The controls available on the EVOS M5000 Imaging System are ™...
Appendix B: Graphical user interface (GUI) Capture tab Viewing area: Displays the sample. Zoom slider: Zooms in and out of the Viewing area. The zoom range is 100% to 1000%. Active objective and phase ring: Displays the active objective and phase ring information. Channel: Selects the light source from the installed LED light cubes (fluorescent channels) or from the condenser (transmitted light).
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Brightness mode: Toggles between Simple and Actual modes for brightness controls. • Simple mode allows you to control light intensity as a single Light parameter. • Actual mode allows you to adjust Brightness (i.e., LED intensity), Exposure, and Gain parameters individually.
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Display settings and analysis tools: Allow you to change image display settings in the Viewing area, and analyze and annotate captured images. Image Display Settings: Opens the Image display settings tool, which allows you to adjust image display parameters (Brightness , Constrast , Gamma Correction ) for the selected...
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Show Cell Count: Allows you to perform cell counts using the Auto Count or Manual Count tools. • Auto Count: Allows you define auto count parameters by selecting representative target objects, then count the objects by intensity, area, and circularity. •...
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Z-Stack: Opens the Z-Stack tool, which allows you to capture multiple images along the Z-axis based on your specifications. You can use the images captured with the Z-Stack tool to generate a Z-Stack Projection. Save: Opens the Save dialog, which allows you to select a save location, set save options, and enable the Quick Save option.
Review tab Viewing area: Displays the sample. Folder: Displays the location of the current folder or image. Up: Goes up one folder from the current folder. Refresh: Refreshes the list or grid of images in the current folder. Grid view and List view buttons: Display the folders or saved images in grid or list view. Folder/Image display size: Increase and decrease the display size of the folders or image files.
Settings Objectives: Allows you to assign and unassign objectives on the objective turret, and to calibrate objective magnification. Visuals: Allows you to calibrate color channel lighting (White Balance Calibration) and to assign saturated pixel colors in the Fluorescence, Transmitted, and RGB Transmitted channels.
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Filter Cubes: Allows you to add or remove EVOS ™ light cubes from the instrument and to assign pseudocolors to specific channels. Network: Allows you to connect to a Wi-Fi network and to map network drives. Service: Displays the EVOS M5000 hardware and ™...
Appendix C: System overview Technical specifications Note: Technical specifications of the EVOS ™ M5000 Imaging System are subject to change without notice. For the latest product information, see the product page (thermofisher.com/evos). Physical Dimensions (H × D × W): 46 × 59 × 46 cm (18 × 23 × 18 in) characteristics Weight: 16 kg (35 lbs.) Footprint: Approximately 92 cm ×...
Operation principles and technical overview LED illumination The EVOS ™ M5000 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.
Appendix D: Safety Safety conventions used in this document Safety alert words Three safety alert words appear in this document at points where you need to be aware of relevant hazards. Each alert word—CAUTION, WARNING, DANGER— implies a particular level of observation or action, as defined below: CAUTION! –...
Symbols on instruments Electrical symbols The following table describes the electrical symbols that may be displayed on Thermo Fisher Scientific instruments. on instruments Symbol 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.
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Safety symbols The following table describes the safety symbols that may be displayed on Thermo Fisher Scientific instruments. Each symbol may appear by itself or in combination with text that explains 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.
Safety labels on instruments The following CAUTION, WARNING, and DANGER statements may be displayed on Thermo Fisher Scientific instruments in combination with the safety symbols described in the preceding section. Hazard English Français Symbol CAUTION! Hazardous chemicals. Read the ATTENTION! Produits chimiques dangereux. Safety Data Sheets (SDSs) before handling.
General instrument safety WARNING! PHYSICAL INJURY HAZARD. Use this product only as specified in this document. Using this instrument in a manner not specified by Thermo Fisher Scientific may result in personal injury or damage to the instrument. Moving and lifting CAUTION! PHYSICAL INJURY HAZARD The instrument is to be moved and the instrument...
Chemical safety Chemical hazard WARNING! CHEMICAL HAZARD. Before handling any chemicals, refer to warning the Safety Data Sheet (SDS) provided by the manufacturer, and observe all relevant precautions. WARNING! CHEMICAL HAZARD. All 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.
Chemical waste safety Chemical waste CAUTION! HAZARDOUS WASTE. Refer to Safety Data Sheets (SDSs) and hazard local regulations for handling and disposal. Chemical waste To minimize the hazards of chemical waste: safety guidelines • 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.
Electrical safety DANGER! ELECTRICAL SHOCK HAZARD. Severe electrical shock can result from operating the EVOS ™ M5000 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. Fuses WARNING! FIRE HAZARD.
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 WARNING! BIOHAZARD. Biological samples such as tissues, body fluids, and blood of humans and other animals have the potential to transmit infectious diseases.
Safety and electromagnetic compatibility (EMC) standards This section provides information on: • U.S. and Canadian safety standards • European safety and EMC standards • Australian EMC standards U.S. and Canadian The CSA C/US Mark signifies that the product meets applicable U.S. and Canadian standards, including those from CSA, CSA America, ANSI, ASME, safety standards ASSE, ASTM, NSF and UL.
Documentation and support Related The publication numbers in this section are for the latest product versions available at the time of publication. For documentation supporting newer product documentation versions, go to thermofisher.com/support. Document Pub. No. EVOS M5000 Imaging System Quick Reference Guide MAN0017565 ™...
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thermofisher.com/support | thermofisher.com/askaquestion thermofisher.com 07 Aug 2018...
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