Cytometry and Antibody Technology

Instrument Guides

Work in progress
 
Quick Guides

 

Attune Cell Concentration Recommendations

Spectrum Viewers
 
Instrument Shutdown Protocols for Long-Term Storage
Amnis/Luminex ImageStream

Download Here (With images)

Initializing in 70% isopropanol

  1. Run the Shutdown script
  2. After sterilization completes, remove Sheath bottle, empty PBS and replace with 70% isopropanol (70% IPA)
  3. Place Sheath bottle with 70% IPA back into the instrument
  4. For MKII only: Remove bead vial and empty remaining beads
  5. For MKII only: Place 70% IPA into bead vial and put the vial back onto the system
  6. Run Start Up script
    • For MKII only: uncheck the box “Run ASSIST after Initialization”
  7.  After instrument has finished initializing, stop fluidics
    • note: core will not form because there are no beads and the sheath is 70% IPA

Removing excess 70% IPA from the system

  1. Remove all liquid bottles (except Waste bottle) from the instrument, then empty and rinse
    • Sheath, rinse, debubbler, cleanser, sterilizer, and bead vial (MKII only) (do NOT remove Waste Bottle)
  1. Place paper towels underneath all fluidic lines that were previously in bottles
  2. Go to Fluidics tab in Advanced Controls
  3. On the Sheath Pump, change Velocity to 150
  4. Place Sheath Valve to Rinse
  5. Click Empty on the Sheath Valve (Velocity Empty, not Relative Empty)
  6. Observe liquid pouring out Rinse bottle fluidic line
  7. Click Stop on Sheath Pump after 5 seconds
  8. Change Sheath Valve to Reagent
    • Change Sample Flush Valve to Open
    • Change Core Valve to Open
  9. Change Sample Valve to Cleanser
  10. Click Empty on Sheath Pump, observe liquid pouring out of Cleanser line
  11. Click Stop on Sheath Pump after 5 seconds
  12. Change Sample Valve to Debubbler
  13. Click Empty on Sheath Pump, observe liquid pouring out of Debubbler line
  14. Click Stop on Sheath Pump after 5 seconds
  15. Change Sample Valve to Sterilizer
  16. Click Empty on Sheath Pump, observe liquid pouring out of Sterilizer line
  1. Click Stop on Sheath Pump after 5 seconds
  2. Change Velocity to 400 on Sheath Pump
  3. Change Sheath Valve to Vent
  4. Click Empty on Sheath Pump and empty the syringe of all liquid
  5. Change Flush Valve to Rinse
  6. Change Flush Pump Velocity to 150
  7. Click Empty on Flush Pump
  8. Click Stop on Flush Pump after 5 seconds
  9. Change Flush Valve to Flush
  10. Click Empty on Flush Pump
  11. Click Stop on Flush Pump after 5 seconds
  12. Change Flush Valve to Vent
  13. Change Flush Pump Velocity to 400
  14. Click Empty on Flush Pump and remove all liquid from syringe
  15. For MKII only: Change Bead Valve to Flush
    • Change Bead Selector Valve to Waste
    • Change Bead Pump Velocity to 40
    • Click Empty on Bead Pump and remove all liquid from syringe
  16. All syringes should be empty at this point
  17. Next, ensure all valves and vent solenoids are in the “Blue” position
    • Empty and clean Waste Bottle
  18. Place all clean, empty and dry bottles back onto instrument
    • Waste, Rinse, Sheath, Cleanser, Debubbler, Sterilizer, Bead Vial

Additional Note
When the instrument is ready to be restarted, fill the tanks wit the appropriate fluids and run The Shutdown Protocol twice. Following that, Run the Startup procedure as normal.

BD Accuri C6

Cleaning and Decontamination – Standard Procedure

This procedure should be performed when preparing the system for storage or shipping it for service. Contact Accuri Technical Support for more information on shipping. Visit www.AccuriCytometers.com for information on service contracts.

Take appropriate safety measures such as wearing gloves and proper laboratory attire while performing this procedure.

  1. Turn on the Cytometer and open the BD Accuri C6 Software. BD Accuri C6 Flow Cytometer Instrument Manual 7820018 Rev-2 25
  2. Place a sample tube containing 3 mls of diluted Decontamination Fluid (PN 653154) on the SIP and run the instrument for 5 minutes on FAST.
  3. Place a sample tube containing 3 mls of 0.22 µm filtered, DI water on the SIP and run the instrument for 2 minutes on FAST.
  4. From the Instrument menu in BD Accuri C6 Software, activate the manual decontamination fluid cycle by selecting “Run Decontamination Fluid Cycle”.
  5. When the Decontamination Fluid Cycle is complete, turn off the BD Accuri C6
  6. Remove tube of 0.22 µm filtered, DI water from SIP.
  7. Disconnect the fluidic harness from the C6 and the fluid bottles, empty the four fluid bottles, and wipe down the outside of the instrument with 70% ethanol.
  8. Pack the C6, fluid bottles and tray, fluidic harness, and power supply in the Pelican Case.
  • When disconnecting the fluidic harness from the cytometer, always disconnect from the bottle end of the harness first to prevent fluid leaks.
  • NOTE: Contact Accuri Technical Support if you are having difficulty performing this procedure due to instrument malfunction, or you need further guidance on the process.

 

BD Arias

Run a normal Shutdown Protocol with ETOH 70%.

BD Fortessas

LONG TERM SHUTDOWN FOR LSR/FORTESSA/X- 20/CELESTA/SYMPHONY OR CALIBUR USERS
Follow standard daily cleaning procedures before continuing.
1) Bypass the sheath filter and fill the sheath container with 70% ethanol or filtered DI water (Make sure that the sheath tank is clean! You can rinse the tank with 10% bleach solution. Be sure to swish the tank around and upside down while fully closed to get even coverage of bleach. Let this sit for 10 minutes before rinsing with filtered DI water and before filling with DI water or Ethanol).
2) Rinse SIT with DI water for 30 seconds in RUN/HI, then move the arm over for 15 seconds.
3) Place 3ml of 70% ethanol solution on the Sample Injection Port (SIP) and place in run on HI for 10 minutes.
4) Move the arm over for 1 minute or until remaining solution is aspirated. Repeat this again using 3ml of DI water w/ arm over.
5) Place system in Run for 30 minutes in HI w/ 3ml DI water on SIT.
6) Place the system in Standby and turn off the power to the cytometer and computer.
7) Place a sign on the cytometer or computer monitors that the system is in DI water or Ethanol. This is to help remind everyone that the system is not ready to run.
8) To take the cytometer out of the shutdown solution, put the sheath filter back in place, empty the sheath tank and fill with sheath. Place a tube with 3ml of sheath on the SIP and run in HI for 30 minutes.

BD LSR-II HTS

Source with pictures here. This is likely applicable to the Fortessa HTS as well; they’re very similar.

 

Placing the HTS for BD LSR II in Long-Term Storage

  1. Remove the HTS safety cover.
  2. Locate and loosen the positioning screw that secures the HTS to the cytometer support bracket (Figure 4-17 on page 133).
  3. Grasp the sides of the HTS unit and slide it forward until it meets the stop in the cytometer support bracket.
  4. Lift the HTS unit off of the cytometer support bracket by doing the following:
    1. Slightly lift the HTS unit to clear the positioning screw.
    2. Being careful not to strain the sheath tubing, slide the unit towards you until you can access the filter. This allows you better access to the connections on the back of the unit.
  5. Detach the sheath line from Sheath (B) port on the back of the HTS unit (Figure 4-18 on page 134). Press the metal button to release the connector. Leave the line attached to the cytometer interface panel.
  6. Connect the purging assembly line to the Sheath (B) port. The purging assembly line is located in the spares kit.
  7. Put the end of the purging assembly line into a 500-mL beaker containing DI water.
  8. Put the safety cover on the HTS.
  9. Choose HTS > Prime; repeat nine times. Priming will replace the sheath fluid with DI water.
  10. Remove the end of the purging assembly line from the DI water and lay it on the benchtop.
  11. Remove the purging assembly line and reconnect the HTS tubing to the Sheath (B) port.
  12. Detach the HTS from the cytometer by following the steps in Removing the HTS Unit on page 159.
  13. Remove the HTS base plate:
    1. Unscrew the two thumb screws on each side of the base plate.
    2. Pull the base plate toward you until it disengages from the cytometer support bracket.
    3. Store the base plate with the HTS unit. 
Bio-Rad S3 / S3e

Source pp 105-110 with figures

Decontamination

The Decontaminate tool is accessible by administrators and can be used to completely decontaminate the system. All materials within the system are compatible with 1% household bleach (580 ppm active chlorine).

It is recommended to run the decontaminate procedure at least once every six months but can be run as often as monthly. This tool should also be used if there is a noticeably high background level in the acquired data. The source could be within the fluidic path. Bacteria or fungus can potentially grow in the lines if samples are not handled using basic cellular sterile technique. The bulk fluidics can also contribute to contamination, despite having internal filters built into the lines.

To run the Decontaminate tool:

  1. Move the loading stage into the wash position.
  2. On the Administrator tab select Decontaminate.

A warning window will appear to confirm that a decontamination procedure should be run.

A decontamination wizard will start and walk through each procedure. When prompted follow the instructions.

  1. Choose whether the system should be shut down after decontamination has completed (Figure 129).
  2. Click the checkmark button to proceed.
  3. Select whether or not an automatic startup should be scheduled (Figure 129).
  1. If yes, assign date and time for automatic startup procedure to run.
  2. Click the checkmark button to proceed.
  1. Replace the DI water and sheath fluid containers with at least 1 L of decontamination solution (Figure 130).
  2. Empty waste container and replace.

10. Click the checkmark button to proceed.

The system will drain the internal sheath reservoirs into the waste container (Figure 131). The system will directly prime the fluidics with the decontamination fluid (Figure 132).

In order to decontaminate, the system will soak the fluidic lines in the decontamination fluid and then will shut down (Figure 134).

After the system shuts down, the containers can be removed and the sheath and DI water cap assemblies should be rinsed with DI water. Sheath fluid and DI water containers can be returned to proper position (Figure 135). The system will drain the sheath fluid reservoir and the sheath filter before priming the system with 8x sheath fluid or DI water (Figures 136–137). After decontamination the system should be ready for running samples. Any high background should be eliminated.

Note: The user must be present to change the containers approximately 1 hour into the procedure. After this point the system can be left overnight to complete the rinse and shutdown if that option is chosen.

The system will soak and go through an extended warmup (Figure 138) before a system startup is commenced (Figure 139).

After decontamination the system should be ready for running samples. Any high background should be eliminated.

•Below is the recommendation to prepare the S3e for long-term storage of 1 to 3 months, where the unit is not being moved from its location:
1.Refer to the S3e user guide for cleaning to ensure the S3e line is clear of cellular debris:
–See ‘Cleaning the system’ on page 102, using the approved disinfectant from page 175 for the sample line (filtered 10% bleach with a high pressure clean, followed by 5 tubes of filtered DI water, with high pressure clean)
2.Open the fluidics status icon shown below, and select the hot swap function.
3.Disconnect the 8x sheath bottle from the line, and remove the bottle to allow it to run dry.  (The timer on the fluidics status icon will indicate when the S3e is out of 8x sheath.)
4.Once the 8X sheath line has run dry, attach a DI water tank to the 8X sheath line position.  If the stream stopped earlier, it should restart once you refill the 8X line with the DI water tank.  (If the stream doesn’t restart as expected, simply run startup again.)  Then run the stream for an hour with DI water to remove all traces of 8X sheath from the fluidics line.  
5.Finally, shut the instrument down (see procedure on page 104 of S3e user guide).  Open the nozzle collar a half-turn. 
6.Gently wipe the nozzle tip once with a Kim wipe to remove any droplets at the end of the nozzle.  This will prevent a serious clog from forming after shutdown.
7.You can now leave the S3e on after shutdown, for short-term storage.
8.If you are concerned about power outages due to the shutdown at your institution for quarantine, then you can completely power off the instrument and computer, and unplug the S3e and PC from the wall outlet.
9.After a few weeks or months of the S3e sitting idle, the nozzle will need to be cleaned and a swap tip performed, before the S3e can be used.  Also remember to replace the 8X sheath tank to its position before using the S3e. 
Bio-Rad ZE5

Source p.325-333 with figures

Decontaminating the System

Note: Bio-Rad strongly recommends that your preventive maintenance program include decontamination of the entire ZE5 Cell Analyzer fluidics system.

You can use the Decontamination wizard to completely decontaminate the system using 1% filtered bleach solution (25 ml household bleach and 2.475 L filtered water). See Cleaning Solutions on page 321 for details. Regular system decontamination is recommended to ensure that lines, bottles, and valves are free of microbial growth.

Use the Decontamination wizard at least once every six months, or as often as necessary. Use this wizard if there is a noticeably high background level of particles in the acquired data. The source could be within the fluidics path. Bacteria or fungi can grow in the lines if samples are not handled using basic cellular sterile techniques. The bulk fluidics can also contribute to contamination, despite having internal filters built into the lines. To test for contamination, disconnect the waste bottle cap, collect fluid in the waste input line, and culture it.

The wizard guides you through the necessary steps to complete the decontamination process, which takes about 4 hr. When the process completes, the system is ready to run samples, although you can run the QC process afterward.

Important: While decontamination is in process, the following functions are disabled:

Returning to the Acquisition window
Viewing the optical filter configuration
In the Instrument tools:

o Returning the sample probe to home position
o Unclogging the sample line, probe, and flow cell o Cleaning the sample line and probe
o Pausing the sheath fluid and disabling the lasers o Swapping QC beads

Opening the sample loader door
Running the QC process
Sending the run list to the local instrument
Shutting down the instrument
Closing Everest software

The system displays a message indicating that decontamination is in progress.

Required Materials

To complete the decontamination process you will need the following materials. n Four extra sheath bottles
Two extra additive bottles
Two extra cleaner bottles

Two standard fluorescence-activated cell sorting (FACS) tubes (12 x 75 mm, 5.0 ml) n 12 L filtered DI water
100 ml filtered bleach (filtered through a 0.2 μl filter flask or syringe filter)

Preparing for the Decontamination Process
Important: When handling sheath fluid and DI water bottles, always wear gloves and minimize air

exposure to help avoid contamination.
– Before beginning the decontamination process you must
– Decontaminate the outside of the probe with 10% bleach solution.
– Prepare bleach and rinse bottles for decontamination.
– Replace both sheath bottles with bottles filled with 2.5 L diluted 1% bleach.

Replace the additive and cleaner bottles with bottles filled with 200 ml diluted 1% bleach.

This section explains how to decontaminate the probe and prepare the bleach solution.

Tip: You can prepare the bottles while the system is decontaminating the probe.
To decontaminate the probe and sample lines

  1. Prepare 100 ml of filtered bleach solution using a 0.2 μl filter flask or syringe filter.
  2. Prepare 10 ml of 10% vol bleach solution in filtered water by adding 1 ml filtered bleach to 9ml filtered DI water.
  3. Fill a stat tube with 3.5 ml of the 10% filtered bleach solution and insert it into the stat tube position.
  4. On the Home window of Everest Software, click STAT Tube to start acquisition.
  5. Run the stat tube acquisition with the filtered bleach solution for 5 min at 1 μl/sec.
  6. After 5 min, stop the stat tube acquisition and remove the stat tube.
  7. Insert a stat tube containing 4.0 ml of DI water into the stat tube position.
  8. Run the stat tube acquisition with DI water for 5 min at 1 μl/sec to remove any bleach from the outside of the probe.

To prepare the bleach bottles for decontamination

  1. Obtain the following materials: n Two 4 L sheath bottles
    – One additive bottle
    – One cleaner bottle- 0.2 μm filtered bleach- Filtered DI water
  2. Prepare each 4 L sheath bottle:
    1. Add 25 ml of filtered bleach solution to each bottle.
    2. Add 2.475 L of filtered DI water to each bottle.
    3. Ensure that the liquid solution in each bottle is thoroughly mixed.
  3. Prepare the additive and cleaner bottles:
    1. Add 2 ml of filtered bleach to each bottle.
    2. Add 198 ml of filtered DI water to each bottle.
 

 

Ensure that the liquid solution in each bottle is thoroughly mixed.

To prepare the rinse bottles for decontamination

  1. Obtain the following materials: n Two 4 L sheath bottles
    – One additive bottle
    – One cleaner bottle- Filtered DI water
  2. Add 2.5 L filtered DI water to each 4 L sheath bottle .
  3. Add 200 ml filtered DI water to the additve bottle and the cleaner bottle.

To prepare the system for decontamination

– In the instrument’s fluidics chamber, replace both sheath bottles and the additive and cleaner bottles with those containing the filtered bleach solution.

See Replacing Sheath Bottles on page 146 and Replacing Sheath Additive and Cleaner Bottles on page 148 for information about removing and replacing the bottles.

Important: The decontamination process takes approximately 4 hr to complete. During the process, you will be prompted to

– Replace both sheath bottles approximately 50 min into the procedure with sheath bottles filled with 2.5 L of DI water.

– Replace the additive and cleaner bottles filled with 200 ml of DI water.
– Replace both sheath bottles again approximately 190 min into the procedure with the default (original) sheath bottles filled to the Fill Line with sheath fluid.

– Replace the additive and cleaner bottles with the default (original) additive and cleaner bottles filled to the Fill Line with the appropriate solutions.

Before running the wizard, ensure that the waste bottles are empty. The bottles containing the bleach solution must be filled no more than halfway.

Important: When handling sheath fluid and DI water bottles, always wear gloves and minimize air exposure to help avoid contamination.

To decontaminate the system

  1. Before starting the procedure, ensure that you have performed the steps in Preparing for the Decontamination Process on page 326.The Decontamination wizard will prompt you to replace the existing bottles.
  2. In the System section of the toolbar, click Decontamination in the Instrument Tools dropdown list.The Decontamination wizard starts.
  1. Carefully read the contents of the screen. Verify that you have completed the preparation steps and click Continue to proceed with the Decontamination process.Important: If you have not completed the preparation steps, click Cancel to stop the Decontamination wizard and perform the steps in Preparing for the Decontamination Process on page 326 before continuing.
  2. To view the decontamination progress details, click the down arrow on the Decontamination button.
  1. Approximately 50 min into the procedure, the software prompts you to replace both sheath bottles with bottles containing 2.5 L of DI water, and replace the additive and cleaner bottles with bottles containing 200 ml of DI water.
  2. After replacing the bottles, click Continue.
  3. Approximately 190 min into the procedure, the system prompts you to replace both sheath bottles with bottles filled to the Fill Line with sheath fluid, and replace the additive and cleaner bottles with the default (original) bottles filled to the Fill Line with the appropriate solutions.

8. After replacing the bottles, click Complete.

Everest Software initiates a fluidics startup and then a fluidics shutdown to complete the process. This can take an additional 12 min.

After decontamination completes, close the software, power off both the computer and the instrument. If the ZE5 Cell Analyzer is not connected to a UPS (uninterrupted power supply), then unplug it from the wall outlet.

Cytek Aurora

Source

If you need to shut down your Aurora or Northern lights cytometer for an extended period of time (e.g. more than 1 week), follow the steps below to properly clean and decontaminate the fluidic lines to ensure the system will operate smoothly when it is time to turn it back on.

Materials

  • Approximately 1L of 10% bleach (10% is diluted Clorox household bleach, which is around 6.0% sodium hypochlorite)
  • Approximately 2L of DI water – this is key to prevent bacterial growth in the lines while the system is off
  • Two 5mL polystyrene tubes containing 3mL 10% bleach
  • Four 5mL polystyrene tubes containing 3mL DI water

Steps

  • While the instrument is on and connected to SpectroFlo, go to the Cytometer Menu.
  • If time permits, perform an overnight Contrad soak to thoroughly clean the flow cell by running Fluidics Shutdown with 2 modifications:
    • In Step 4, load another tube containing 3mL 25-50% Contrad.
    • Once Fluidics Shutdown is completed, remove the Contrad tube from the cytometer and load a tube containing approximately 1mL DI water.
    • Power off the system and leave it overnight.
    • The next morning, power on the instrument and connect to SpectroFlo.
  • Run a Long Clean. Follow the steps in the wizard to complete the Long Clean (video tutorial here, skip to minute 3:18).
  • When the Long Clean has completed, run Fluidics Shutdown. Instead of running the tubes prompted by the software wizard (10% bleach, DI, 30% contrad, DI), ONLY run DI water tubes.
  • When the Fluidics Shutdown is complete, power off the instrument and leave a tube containing approximately 1mL DI water on the sample injection port (SIP).  This will keep the sample line wetted and prevent it from drying out.

Resuming Operation

  • When you are ready to use the instrument again, power on the instrument.  
  • Open SpectroFlo, log in, and wait for the software to connect.
  • Optional: If you’d like to replace the DI water in the system with a different sheath, run a Long Clean using 10% bleach, and then your desired sheath.
  • Open a Default Experiment and set flow rate to Medium.
  • Load a tube containing approximately 2mL DI water and click start.
  • After 30 minutes, click Stop.
  • Run QC to ensure the cytometer is functioning properly.
Coulter CytoFLEX

Source from CytoFLEX Manual with Pictures (pp. 11-14 to 11-17)

Preparing the Instrument for Transport or Storage

When the instrument is to be transported or is not to be used for 30 days or more, complete the emptying processes to prevent instrument damage and to reduce the possibility of biological contamination. Contact us if you have any questions.

  1. Run the Deep Clean procedure. (Manual pp. 11-9)
  2. Run the Daily Clean procedure. (Manual pp. 11-1)
  3. Clean the Sample Station. (Manual pp. 11-7)
  4. Empty the sheath fluid container and the waste container. (Manual pp. 12-9)
  5. Clean and disinfect all surfaces. (Manual pp. 11-13)
  6. Clean the sheath fluid container and the waste container. (Manual pp. 11-10)
  7. Remove the right-side cover. (Manual pp. 12-4)
  8. Remove the Deep Clean solution bottle from the bracket, empty the Deep Clean solution bottle, and rinse with DI water. Then, attach the Deep Clean solution bottle to the bracket.
  9. Remove the Plate Loader module if applicable. (Manual pp. 12-51)
  10. Power down and disconnect all the cables and sheath fluid and waste harnesses. 11-16 B49006AP Cleaning Procedures Nonscheduled Cleaning
    1. CAUTION Risk of instrument damage. The Cytometer can suffer irreparable damage if it is exposed to subfreezing temperatures while it still contains liquid. Always drain the flow cell after cleaning the Cytometer if the Cytometer will be stored or transported in subfreezing temperatures.
    2. WARNING Risk of contamination from biohazardous material. Always wear PPE when performing this procedure as you can contact components with blood residue when handling the Fluidics module. Dispose of any absorbent materials used to collect or clean up leaks in accordance with the local regulations and acceptable laboratory procedures.
  11. Disconnect the blue-labeled tubing from the pneumatic valve PV1 and hold the absorbent material under the disconnected tubing to collect any dripping liquid.
  12. Disconnect the yellow-labeled tubing connected to the choke to vent the flow cell, allowing it to drain.
  13. Verify that liquid has stopped dripping from the blue-labeled tubing.
    1. NOTE The flow cell is empty when liquid stops dripping from the blue-labeled tubing.
  14. Dispose of the absorbent material used to collect the liquid in accordance with the local regulations and acceptable laboratory procedures and cleanup any spills.
  15. Reconnect the blue-labeled tubing to PV1.
  16. Reconnect the yellow-labeled tubing to the choke.
  17. Reinstall the right-side cover. (Manual pp. 12-4)
  18. Ensure that the optical filters are seated properly.
  19. Ensure that the top cover is tightly closed. 
Coulter MoFlo Astrios

Given the lack of a specific long-term storage procedure for the Astrios, Beckman Coulter’s lead engineer has recommended a Yearly Decontamination.

Source from Astrious Manuel with Pictures (pp 9-28 to 9-32)

Yearly Fluidics System Decontamination

  1. Obtain and put on required personal protective equipment which includes lab coat, gloves and safety glasses.
  2. Obtain the following supplies:
    1. One new in-line sheath filter.
    2. A bleach solution containing 2000 ppm active chlorine is required (115 mL household bleach + 2875 mL water). Use only high-quality, fragrance-free, gel-free bleach (5 to 6% solution of sodium hypochlorite – available chlorine).
    3. Paper towels
    4. Large bucket
  3. Press the Change Tanks button to depressurize the system.
  4. Remove and discard the in-line sheath filter. Reconnect the canister without a sheath filter installed. Examine the canister for contamination, clean as necessary.
  5. Place 3 L of decontamination solution into an empty sheath tank and connect to instrument. Prepare a 5 mL tube of decontamination solution for placement on the SmartSampler.
  6. Press the Start Fluidics button.
  7. While the smart sampler is running, select fluidics on and off 5 times and rinse probe at least 2 times.
  8. Disconnect the nozzle assembly connector from the left cover panel.
  9. Turn the knob on the nozzle stage and raise the nozzle.
  10. Loosen the knob on the front of the nozzle body and then pull the nozzle body out from the stage.
  11. Turn the nozzle body upside-down, unscrew the black retaining ring, and remove the nozzle tip.
  12. Locate a large bucket in which to drain the nozzle.
  13. Start fluidics and run the 3 L of decontamination solution into the bucket.
  14. When the sheath tank is empty, press the Change Tanks button . IMPORTANT The tanks and filter canister must be thoroughly rinsed.
  15. Remove the sheath and waste tanks as well as the sheath filter canister. Rinse the tanks and the canister with deionized water.
  16. Fill the sheath tank with deionized water.
  17. Return the tanks and canister to the instrument.
  18. Start fluidics and let the tank drain into the bucket.
  19. Repeat Steps 14 – 18 for another tank of deionized water.
  20. Using sterile technique, install a sterile sheath filter in the canister.
  21. Fill the sheath tank with deionized water and run fluidics to let the tank drain into the bucket. Debubble using the Sheath Filter Vent Lever frequently. NOTE Repeat this step two times so that two tanks of deionized water have run through the new sheath filter.
  22. Fill the sheath tank with sheath. WARNING Risk of personal injury. Be careful not to touch the exposed injection tube. When reinstalling the nozzle tip, the exposed injection tube could puncture your skin.
  23. Reattach the nozzle tip.
  24. Reinstall the nozzle body in the stage. WARNING Risk of personal injury. When lowering the nozzle stage, you could pinch your fingers between the bottom of the stage and the instrument frame. Lower the nozzle stage using the upper portion of the stage to avoid pinching points.
  25. Lower the nozzle stage back into the instrument and lock it in place.
  26. Reconnect the Nozzle Assembly connector.
  27. Debubble for one hour by pressing the Debubble button .
  28. Check the nozzle and fluidics lines for visible bubbles and debubble again if necessary
Fluidigm Helios

Extended cleaning prior to Helios Shutdown

  1. Clean the sample probe line, sample line, sample capillary and nebulizer by running wash solution for at least 5 min with wash solution and ultrapure water (DIW) for at least 10 min.
    • Optional: Wash solution can be run for longer than 5 min if desired. However, make sure to rinse with DIW for at least 2 times as long as the wash solution is run.      
    • Note: If plasma is not on, follow steps 1-8 for cleaning procedure without plasma in the Helios Users Guide (page 76). 
  1. Remove the sample capillary from the nebulizer.
  2. Remove the nebulizer from the nebulizer adaptor.
  3. Disconnect the neb gas line from nebulizer.
  4. Clean the nebulizer according to the nebulizer cleaning procedure as described in the Helios Users Guide (pages 93-94).
  5. Store the nebulizer by soaking in DIW in a covered container.
  6. Disconnect the argon gas line from the sample loader.

Helios Shutdown

Follow the instrument shutdown procedure provided in the Helios Users Guide (page 80 )

 Important NoteIf you decide to shut off the Argon supply to the instrument, you will need more time to recover the vacuum levels in the TOF chamber when the time comes to start-up the instrument. This also applies if you choose to leave the Argon supply on and eventually run out of Argon – the system will need more time to recover the vacuum levels. 

 Helios Startup

Please follow the start-up procedure provided in the Helios User Guide  (page 81)

Important NoteIf the Argon supply was shut off, start the Argon gas supply for two hours prior to re-starting the Helios.

Recommendations for Data Backup for both Helios and Hyperion Systems

We recommend that you backup your data as well as the CyTOF ‘conf’ folder following the steps below.

  1. Backup all the data from the E drive to an external and/or Network drive. Please note that we highly recommend backing up your data after each run. If you are not sure about the size of the data, which exists on the E drive, please ask your IT department for advice about the size of the target backup drive.
  2. Close the CyTOF software and wait for 10 seconds.
  3. Open the file explorer and navigate to C:\ProgramData\Fluidigm\CyToF
  4. After verifying that the software is closed, make a copy of the “conf” folder onto the same external drive and/or the Network driver where you have backed up your sample generated data.
Luminex 100/200

Storing the System

This procedure explains the steps you should take before placing the system into long-term storage.

  1. Run a sanitize with 10% to 20% household bleach solution.

  2. Run a sanitize with distilled water.

  3. Run four washes with distilled water.

  4. Remove the sample probe from the instrument, flush with distilled water from the narrow end out through the larger end, replace it in the sample arm, and wrap the end with Parafilm® M.

Miltenyi MACSQuant Analyzer

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Long term storage of MACSQuant® Analyzer

Prepare your MACSQuant® Analyzer for periods of non-usage

If you need to shut down your MACSQuant for a longer period of time, follow the instructions below to clean the fluidics of your MACSQuant and to ensure that the instrument will run smoothly when turning it back on.

  1. Run a Clean program by right-clicking the Rinse button and selecting Clean. Follow the on-screen instructions:
    1. Add 0.5 mL of a 1% hypochlorite solution into a 5 mL tube.
    2. Click Continue.
    3. The cleaning process will take approximately 10 min.
  2. Run a Flush program by right-clicking the Rinse button and selecting Flush. The process will take about 15 min.
  3. Shut down the system manually via the Main instrument control button .
    For further details on these procedures, please refer to the MACSQuant instrument manual or the short instructions “Start up and shutdown”.

What to consider during the periods of non-usage

In order to prevent clogging of your MACSQuant® Analyzer 10, 16, X or VYB over time, the instrument should be run at least every two weeks. To this end, turn on the instrument and switch it into acquisition mode at least every two weeks. During the priming procedure, the fluidics will be flushed and any potential residues will be removed. After the instrument has been primed and all system checks have been performed, shut down the instrument manually via the Main instrument control button . For further details on these procedures, please refer to the MACSQuant instrument manual or the short instructions “Start up and shutdown”.

In some cases, it might be required to store your MACSQuant Analyzer for a prolonged period of time without access to the instrument at least every two weeks. In this instance, please start a MACSQuant Live Support session, contact your local technical support or your local MACSQuant Specialist for further details on how your MACSQuant may be prepared for a long- term storage of > two weeks.

 

Sony SH800 Cell Sorter

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SH800 Cell Sorter

Long term shutdown (4 weeks or greater)

  1. Power up the instrument and the SH800 Software
  2. At the software log-in screen, select the ‘Maintenance’ tab in the bottom right corner. For SH800Z and SH800S models select “Ethanol Cleaning of All Lines.”

NOTE: For SH800Z models without the “Ethanol Cleaning of All Lines” option- follow the manual cleaning procedure using sterile DI water instead of 70% ethanol

  1. Proceed with the wizard to fill the system with 70% ethanol and select the 60-min soaking option.
  2. At the rinse step of the wizard, when the software says to fill the sheath tank with PBS, use 3L of sterile DI water instead. This way the lines will be filled with DI water for long term storage.
  3. Once the cleaning finishes, power down the system and the workstation. Empty the Waste and Sheath tanks on the fluidics cart and the DI water bottle behind the left side door.

Starting Up After Long Term Shutdown

  1. Power up the instrument and the SH800 Software
  2. At the software log-in screen, select the ‘Maintenance’ tab in the bottom right corner. For SH800Z and SH800S models select “Ethanol Cleaning of All Lines.”

NOTE: For SH800Z models without the “Ethanol Cleaning of All Lines” option- follow the manual cleaning procedure.

  1. Fill the sheath tank with 3L of sterile water, and the DI water tank with 500mL of DI water
  2. Click ‘Ethanol Cleaning of All Lines’ in Maintenance tab at the bottom right corner of the log in screen and proceed with the cleaning wizard.
  3. Allow the system to soak overnight with ethanol. At the end of the cleaning cycle, fill sheath tank with fresh sheath fluid when prompted and thoroughly de-bubble the filters.
  4. Start system setup and calibrate the system using sorting chip.

Note – If the stream is unstable, run another ‘Ethanol Cleaning of All Lines’ in Maintenance tab at the bottom right corner of the log in screen cleaning and allow run ethanol cleaning again allowing it to soak the lines for 3-4 hours.

ThermoFisher Attune NxT

Attune Long Term Shutdown with images

Long Term Shutdown

  1. Run Thorough Shutdown with 10% bleach (or better yet flow cell cleaning solution 1:3) to decontaminate Attune NxT.
  2. Or, run a System Decontamination script.  Note: This script will take longer than a Thorough Shutdown and will require two new focusing filters when the instrument is brought back online.
  3. Once the Shutdown or System Decontamination has completed, empty out the Focusing Fluid Container and the Auto Sampler Focusing Fluid container, if present. Plug the empty focusing fluid container on its side to “trick” the floating focusing fluid sensor so that the container to read as full.  If the blue light is still flashing, tip the bottle and tap the bottom so the sensor (see red circle) slides toward the bottle cap.  Repeat for the Auto-Sampler Focusing Fluid container.
  4. Run the Startup script.
  5. Repeat the Startup script until the Filtered Focusing fluid reservoir located in the back-left corner of the Attune NxT (next to the sample syringe) is empty.
  6. Please note that software may repeatedly show Error 13 – Fluidic Errors (buffer tank not refilling) and the startup will stop.  Click OK once the error shows up and repeat the startup script.  This should take approximately 4-5 startups to empty the Filtered Focusing Fluid Reservoir.  This step will remove the PBS (0.1M Sodium phosphate) out of the system so that crystallization should be less of a problem
  7. Once the Filtered Focusing fluid Reservoir is empty, rinse and fill the Focusing Fluid container(s) with 400ml Sterile water if possible or Di-water.  Keep adding water until the floating level sensor triggers.
  8. The Filtered Focusing Fluid Reservoir in the back corner should start filling up.
  9. If the pump does not turn on to fill the reservoir, run the Startup script or Rinse script.  The Filtered Focusing Fluid Reservoir does not need to fill to the top unless you ran a System Decontaminate script, but only a quarter way is enough.
  10. Then disconnect the focusing fluid tank sensor cable to stop the Focusing Fluid Reservoir tank from filling.  This step flushes the remaining focusing fluid out of the instrument which contains PBS and detergents.
  11. Empty out the Focusing Fluid Container(s) and plug upside down as you did in Step 2.
  12. Repeat running Startup script until the Filtered Focusing Fluid Reservoir is empty.  The Attune is now ready for long term storage as the instrument is free of salts and dry.
  13. Power off the Attune NxT and the auto-sampler using the switches located in the back.  Empty the remaining containers.

Turning on Attune NxT after Dry Storage

  • If the instrument was stored after running the Thorough Shut down script, then the Attune can be re-started by filling the fluidics bottles with the appropriate solutions and running Startup 3 times, De-bubble 2 times and Rinse 1 time.

OR

  • If the instrument was stored after running the System Decontamination script, replace the Focusing Filters with new filters, fill containers with appropriate solutions, run Startup 3 times, De-bubble 2 times and Rinse 1 time.