Dr. Amanda Capes-Davis, an independent cell culture consultant and founding manager of CellBank Australia, talks to contributing editor Tanuja Koppal, Ph.D., about the imperative need to establish best practices in a cell culture laboratory. She talks about everything that can be done from the early stages in terms of utilization of lab space to establishment of procedures for minimizing contamination and also planning for emergencies and disaster recovery.
Q: What are some of the key elements that lab managers should consider when setting up a cell culture lab?
A: There are some things a lab manager simply cannot control, and one of them is the space they have for doing cell culture. But if possible, they should try to dedicate a space exclusively for cell culture. If you are working with cells and someone over your shoulder is working with bacteria, then you know that you are going to have problems down the line. The next thing that you should think about is the equipment for cell culture. When you are choosing equipment for a new lab, you have to remember that you are going to be living with those choices for years. So you want to have things that can be cleaned easily and you should know where to get them serviced or calibrated if there is a problem. You should look at the latest technology, especially when it comes to cleaning incubators, and make decisions based on what is the best you can buy within your budget. When you are buying consumables you want to focus on those that are cell culture grade. They may be more expensive, but essentially you are paying for the testing that the supplier has done for the relatively high-risk problems that can occur. I recommend using as many disposables as your budget allows, as it can reduce a lot of problems. Finally, when you obtain your cell lines, make sure that you obtain them from a supplier that also tests the cells, particularly for contamination. You need to know what types of tests have been done on the cells that are coming into your lab. A lot of the choices you make can have an impact on the problems you face later.
Q: What is the biggest source of contamination in a cell culture laboratory?
A: The most common source of contamination today is other cell lines. In past years, consumables were also a big problem, but suppliers these days do the necessary testing so that risk is rare. Contamination from other cell lines often occurs when people share a bottle of medium between cell lines. Some degree of contamination risk also comes from the operator or the donor. Based on the data that I have seen, mycoplasma and cross-contamination are the two most significant causes of contamination. Of course, contamination from bacteria and fungi is also common, but people are more likely to see those organisms under the microscope or they can see changes in the cell culture medium. Whereas changes caused by mycoplasma and cross-contamination are more subtle. If you have been working with the cell line you may see the change, but if you are new to the cell line, then it’s easy to miss seeing the changes.
Q: What do you suggest in terms of keeping the cell culture area clean and contamination-free?
A: Testing for mycoplasma and cross-contamination is very important, and if you test for it regularly you are likely to pick up the problem early. A lot of people do a quick test for mycoplasma every month to make sure there is nothing detectable emerging that is problematic. At the minimum you want to test for mycoplasma and cross-contamination when you put down a stock in liquid nitrogen and you can keep coming back to the stock, say every three months. So you are always starting with something you have tested. Having good aseptic technique and separating the cell cultures so that you are not sharing medium or other consumables is important. Having some separation in time between handling different cell lines in the hood is also very important. For most labs, growing different cell lines in different incubators is not feasible and, hence, it’s reasonable to share incubator space.
Q: What do you recommend in terms of disaster planning?
A: It is always good to assume that any new cultures coming into the lab could cause a problem. If you are starting out in a new lab, it’s a good idea to set aside some space or an incubator to hold new cells before you have a chance to test them. You want to separate these cells from other cultures until the test results are back, particularly to avoid mycoplasma contamination that can spread across different cultures very easily. At a building level, all labs should think about having an emergency power supply to run fridges and freezers that contain irreplaceable samples. At the institute hosting our cell bank we have a generator that is tested regularly and can kick in if there is a power failure, and all our essential equipment is connected to that emergency supply. You also need to think about access to the building so that people can come in during an emergency to top off liquid nitrogen in the tanks that store the cells. Think about what needs to be done if elevators are not working and if you need to access the stairs. Finally think about a separate site where you can put some vials of cell lines that are irreplaceable to you. Either have a cell bank or storage facility store them for you or exchange samples with colleagues who are off-site, to cope with disaster recovery. It’s one of those things that you can be tempted to put off until tomorrow, but it’s best to think about it early in the process.
Q: What are some of the common issues that come up as you consult for various clients?
A: I am sometimes asked for help with problematic cell lines that need short tandem repeats (STR) profiling where it may be useful to seek multiple opinions. I am particularly interested in increasing awareness and educating people about cell culture. Generally people need more awareness of the problems that can come up with cell culture, and they find it particularly convincing when they see me present data on the problems that can come up. There are not enough programs to educate people about best practices. People often learn about cell culture based on what’s being done in their own lab. That can be very good but it can also be that nobody ever learned how to do it using best practices. So they need a resource to go to and learn and bring best practices into their labs, so they know that they are doing cell culture as best as they possibly can.
Q: What can be done to educate and train lab personnel to ensure that best cell culture practices are being inculcated and followed?
A: First of all, each lab should have a cell culture expert who can teach new students and lab personnel. Every lab should have resources, such as a good textbook, that people can refer to for various problems that might crop up. Many undergraduate courses cover cell culture, but for most people it’s a long time before they start using that knowledge in a lab. Training for graduate students and postdoctoral fellows is often missed. Some of the cell banks offer training courses, and this is also an opportunity for societies that have a big investment in cell culture. Online courses are good, but a challenge with cell culture is the need to be hands-on. Learning good aseptic technique is important for doing cell culture well, and it’s an advantage to be able to watch someone actually working with cells to see what can be done differently. However, most of the ideas and the background of why and how things are done can be covered well with an online approach.