What liquid handling robot is the best for sterile cell culture work?Performing live cell culture work on a liquid handling system requires special considerations for sterility. These go above and beyond the aseptic conditions most users require for their liquid handling robots. Selecting a liquid handling robot for sterile cell culture requires the purchaser to consider some key concepts. These requirements must be known and communicated to the vendor when discussing a purchase. Regardless if you will be processing bacterial, yeast, mammalian, or insect cell lines these considerations hold true.
HEPA Air Filtration
Sterile cell work requires a HEPA filtered environment when plates or tubes are only opened. Pathogenic cell lines require a full enclosure around the robot to meet the regulatory needs of our site and country.
For non-pathogenic cell lines it is possible to purchase liquid handlers with built in HEPA filtration. This should be able to provide the same benefits as a positive pressure laminar flow hood. Using a built in HEPA solution will save space in your laboratory as full HEPA enclosures are very large. They can also make maintenance of the system quite challenging.
Pipetting Technology
The liquid handling robot pipette technology is another important consideration when selecting a system for the cultivation of living cells. There are multiple specialty liquid handling options such as the acoustic dispensing designed for ultra low volumes. However, for the all purpose liquid handler, you really need to decide between liquid displacement and air displacement.
Liquid Displacement
Liquid displacement system utilize syringes for each channel filled with system liquid (often 1x PBS or water). Retraction of the syringe plunger aspirates the system liquid through the channel. Depressing the plunger forces system liquid back through the tubing and the channel causing a dispense. There is a concern that liquid displacement systems harbor contaminants such as bacterial or fungal spores in the system liquid. Liquid displacement systems are rarely selected as the liquid handler for sterile cell culture due to concerns about contamination.
Air Displacement
Air displacement systems have a solid plunger mechanism housed in the pipetting channel. Pulling back the plunger creates vacuum to aspirate while plunger depression results in dispense. Air displacement systems use the air in your system. If you followed our above suggestions, system air will be HEPA filtered and clean. Air displacement systems are highly desirable as the liquid handler for sterile cell culture.
Sterile Labware Availability
Enclosing your entire system in a filtration system will do you absolutely no good if the labware you are loading onto the robot isn’t clean. Most manufacturers will have “factory clean” labware that they sell. This usually means it is free from any chemicals or debris from the manufacturing process.
If you have worked in a cell culture lab for any length of time you will know that this is not good enough. Cell culture work usually requires sterility. Sterility is achieved through autoclaving, gas sterilization, or gamma/e-beam sterilization using radioactive isotopes. Make sure that the vendor you are using offers sterile tips, if they don’t then make sure that there is a competitor’s tip that will work on your system that can be provided that way.
All the precautions in the world will not protect your cultures if you are dipping a dirty tip into a clean sterile reservoir of growth media. Don’t even suggest antibiotics, that’s a lazy cover up for bad technique. Do it right the first time.
Tip Ejection
Something a lot of scientists may not think about is that happens to their used tip when they eject it. If you still have liquid in or on the outside of your tips when you go and eject them to your waste you run the risk of creating aerosols which can float about your cabinet. There are many ways to test for aerosolized liquids using fluorescent dyes but it would be far preferable if you didn’t need to worry.
Either having a shielded waste to prevent any aerosolized particles from entering the system deck or an ejection system which uses less force and is therefore less likely to create aerosols would be beneficial. You also should consider the placement of common reagent reservoirs in relation to the waste areas on your system to further reduce this risk.
Loading and Unloading
You now have a nice sterile HEPA enclosed liquid handler and a series of sterile labware, congrats, do you have a plan on how to put those together? Some systems have integrated stackers underneath or on the side, you can purchase a third party incubator or stacker and integrate to your system (assuming you have a gripper arm capable of reaching the hand off location), you can use something like an autoloader, or you can have your users load everything by hand.
Integrated stackers are likely your best solution. Stackers are available off the shelf from the manufacturer of the liquid handler or a 3rd party. They almost always contain barcode scanners which you will likely want on your system for traceability reasons. Stackers and incubators designed to hold plates and reservoirs shouldn’t have any issues accommodating lids to ensure the interior of your labware remains sterile.
Front facing autoloaders like in the Hamilton STAR and Vantage line will work for plates and reservoirs with lids as well. Manual loading could be acceptable but it’ll likely be slower and introduce some additional potential for errors.
Loading tips sterilely is quite difficult if they are not packaged as individual enclosures with lids. A stacker or auto-loader on a system with a large enough external HEPA enclosure that guarantees the tips are in the sterile environment might work.
If having users manually load tips, consider working with a few different ones to determine a protocol. I once wrote a method for loading tips sterile that worked great for me. But my arms are long and when we tried to roll it out to the users the shorter ones were less than enthusiastic. We had to return to the drawing board.
Cleaning
This last point really ties everything together, you need to come up with a cleaning protocol which is suitable to ensure sterility but also is safe for your system.
Many manufacturers will sell systems with UV lights, there is nothing wrong with using UV for an additional safety barrier but do not consider this a sterilization method. UV is great when working with DNA to ensure no cross contamination between samples. If you are relying on UV to kill bacteria or mold, you will be disappointed.
Spray isopropyl alcohol on a lint free rag and wipe down the deck and carriers is the first step. You can also wipe down most arms and channels. Stay away from any motors, threaded rods and gears, and electrical components. Alcohol dries out rubber O-rings so avoid connections with them. Quaternary ammonia compounds may be preferable for some laboratories but the protocol and the logic remains the same. Rule of thumb: If it looks like it shouldn’t get wet then keep it dry.
Your laboratory protocols for your users should have guidelines for when to clean a system. There is nothing wrong with starting your methods with a required series of steps prompting the user clean the system. It is worth it if it prevents one user from spreading a contaminated sample across your entire group.
In Summary
I hope this article helps to make decisions when selecting equipment for automated cell culture system. I’ve found that Hamilton STARs meet most of the requirements on this list. I was able to work around those that didn’t . No one has a perfect system though and you’ll need to decide what compromises you and your cells will be able to live with. If you want to consider other liquid handlers you can also check out Rave Robot’s Liquid Handling Robot List. If you have any questions let us know in the comments below.
