What’s in your cultures?

New recommendations aim to curb cell line contamination

The numbers are alarming, and the costs are great: contaminated cell lines can ruin years of work. Every researcher knows it and, in many cases, has lived through an unwanted discovery. But contaminated lines are pervasive, either coming through the door mislabeled or heterogenous, or unknowingly growing within the lab, the result of mutation or mishandling.

Preventing the use of contaminated or misidentified lines is in everyone’s best interests: the researchers whose careers depend on reproducible, successful trials, the companies and institutions funding the work, and the audiences ultimately benefiting from a new drug or treatment.

If obtaining and maintaining uncontaminated culture lines were easy, however, everyone would be doing it … wouldn’t they? That’s exactly the goal of the Cancer Cell Authentication and Standards Task Force, which has just issued new authentication recommendations presented in the June issue of Nature.

Why cell line authentication isn’t the norm

The task force, formed in 2014 by the Global Biological Standards Institute, was given two tasks:

• “identify and overcome existing barriers to the use of currently available cell authentication tools”
• “support the development, evaluation, and application of policies, novel technologies, and standards for expanded cell authentication”

Theoretically, ensuring an uncontaminated cell line is straightforward. Short tandem repeat (STR) profiling, a type of DNA fingerprinting, can be used effectively to determine the makeup of a cell line.

“The only way to know for sure that your cell line is what you think is to do DNA fingerprinting, by having the test done,” says Mary Kay Bates, global cell culture specialist at Thermo Fisher Scientific.

But knowing that and implementing the process consistently are not the same thing, and researchers may need to do more to identify contamination. Bates adds, “Some people say, ‘I know my cells and I know what they look like,’ but you can’t necessarily tell by looking at them under a microscope.”

Authentication “important but often overlooked”

While staff overconfidence can expose a lab to risk, simple awareness is also a huge issue. Cindy Neeley, senior staff scientist in field applications at Thermo Fisher Scientific, notes, “Cell line authentication, although important, is often overlooked by cell culture researchers for various reasons. It is about time to raise awareness as well as to provide useful information and guidelines to facilitate the practice.”

In looking into the causes for cell line authentication issues, the task force identified no fewer than 10 problems at the root. A sample:

● Prevalence of misidentified cell lines. A 2007 study cited by the task force suggests that 18 to 36 percent of cell lines currently in use are misidentified or cross-contaminated. Moreover, as the Nature piece points out, misidentification can occur when cells are first cultured, when lines are contaminated by unknown cell types, and when cells are mislabeled or contaminated within the lab.
● Authentication of nonhuman cell lines. While many human cells’ fingerprints have been captured, other mammalian identifiers are not as common.
● Naming inconsistencies. A lack of standard naming protocols and syntax for cell lines means the same lines are being labeled differently depending on their source and where they are referenced.

When trust is misplaced

Sharing cell lines is another issue. Bates says, “A lot of it is trust. You get a cell line from a reputable source or you get it from a trusted colleague. But you don’t know that that colleague hasn’t contaminated it themselves.“

In the section on “Acquiring Cell Lines,” the Gibco® Cell Culture Basics Handbook, a free publication from Thermo Fisher Scientific, suggests, “We advise against borrowing cultures from other laboratories because they carry a high risk of contamination.”

The appeal of saving budget dollars can be hard to resist, so the task force suggests that “at a minimum, if a researcher receives a cell line from a colleague, not directly from a reputable cell bank or biomedical research center, then it is imperative to conduct minimal QC to ensure provenance and absence of contamination.”

Bates points out, “While not every lab has the facility to do a DNA fingerprinting test, there are commercial test labs that will do the authentication for you.”

The risks of working by “production line”

Contamination within the lab is another issue, and it can happen to the most experienced researchers. Bates notes, “I was one of those people who was guilty of saying, ‘Oh, I have four cell lines, and they all grow in the same growth medium and the same additives, so I’m going to save time and be a little production line here.’ That’s a terrific way to cross-contaminate your cell lines.

“The one thing that everyone can do—I say this to every group I talk to about cell line contamination—is to only work with one cell type at a time and use different aliquots of growth media for each cell type you work with. Don’t use the same bottle across different cell lines; instead, make up a complete bottle and separate it into smaller portions, one for each cell type.”

According to the task force’s findings, more journals are requiring authentication for submitted research. For that reason and others, authenticating cell lines may yet become part of every researcher’s laboratory practices. Unsurprisingly, the group’s recommendations get to the core of the way laboratory researchers do their work and the culture in which they do it in, advocating for the adoption of standards and best practices, dedicated funding, and a commitment to training, among other changes.

Awareness remains the key. Neeley adds, “Once that level of awareness is up, there might be more interest in developing readily available protocols, methods, and reagents to tackle this issue. Right now, the main issue is that very few people are looking into it, worrying about it, or thinking about it. So we just wanted to bring in some sense of reality.”

Want to learn more about cell culture contamination and the effects it can have on your work? Explore our on-demand webinar: Cell Culture Contamination: What You Don’t Know Can Cost You