Tools for combating cell line contamination and misidentification.

Carolyn Peluso, Ph.D.

Our last blog post told the story of Stanley Gartler who, in 1966, announced to the founding fathers of cell culture that their cell culture collection was contaminated with HeLa cells. He made this stunning discovery incidentally, while in the process of looking for genetic markers to study cancer. He had already used the electrophoretic variance of glucose-6-phosphate dehydrogenase (G6PD) isotypes to demonstrate the clonal nature of cancer in tissue samples (Linder and Gartler, 1965), but wishing to take his research into cell culture, he began characterizing some commonly used cell lines.  What he discovered was that cell lines purportedly representing a wide range demographically, all carried a G6PD isotype specific to the African-American population. The only viable explanation was that HeLa cells had contaminated and taken over many supposedly independent cell lines.

Dr. Gartler, cleverly used the techniques at hand to identify endemic problems in the cell culture community, and then courageously stood up and tried to solve them. Unfortunately, as we also mentioned in our previous blog post, although Dr. Gartler and others tried to eliminate the problems of cell line cross contamination and misidentification, way back in 1966, they remain with us to this day.

Today, however, we have better tools to combat these problems, we just have to make use of them, and below is a list of some available resources to help researchers do just that.

Isoenzyme analysis: Dr. Gartler’s method of using the differences in the electrophoretic banding patterns of isoenzymes is still relevant. There are kits commercially available that provide the necessary reagents to identify the isotype of enzymes, such as aspartate amino transferase or peptidase B, expressed by the cells in question. These kits also provide a comparison chart, so the researcher can determine the species of the cell, and rule out cross-species contamination.

STR profiling: STR profiling is a PCR based approach that can discriminate the origin of the cell line down to the original donor. It is not surprising, therefore, that it is considered the gold standard in cell authentication techniques. Kits containing primer sets are available, but the data is sometimes difficult to analyze without help from a service, such as is available through ATCC. To learn more, please visit the ATCC Cell Authentication Services Page.

ATCC: Your trusted resource

Your trusted resource: The best way to start any project is with material from a trusted resource. We know you trust your buddy in the neighboring lab, but unless you froze the cells yourself, you have no way of knowing what that cryovial holds. What if, for example, the cells were frozen at high-passage number? Or, what if your neighbor’s advisor got them from his neighbor back when he was post-doc! So many variables can only lead to trouble. Cell repositories, like ATCC, on the other hand, will never lead you astray. They check the lines in their collections regularly to ensure that they are properly identified and free from contamination.  Click here for information on the methods used to authenticate the collection of ATCC cell lines.

Hopefully, we’ve helped you start thinking about the best way to authenticate your cell lines. Don’t forget that we are always here to answer questions, and to help in any way we can, so you can move your research forward.

Until next time, when our blog post will focus on helping you choose the most suitable cell line for your experiments, we wish you good luck, and happy culturing,

ATCC

Online resources:

Searchable STR database:

http://www.atcc.org/CulturesandProducts/CellBiology/STRProfileDatabase/tabid/174/Default.aspx

ATCC Cell Authentication Services Page:

http://www.atcc.org/Services/CellAuthenticationTestingService/tabid/1794/Default.aspx

References:

Characterization and authentication of cancer cell lines: an overview. Reid, YA, Methods Mol Biol. 2011; 731: 35-43. Review

Recommendation of short tandem repeat profiling for authentication human cell lines, stem cells, and tissues. Barallon R, Bauer SR, Butler J, Capes-Davis A, Dirks WG, Elmore E, Furtado M, Kline MC, Kohara A, Los GV, MacLeod RA, Master JR, Nardone M, Nardone RM, Nims RW, Price PJ, Reid YA, Shewale J, Sykes G, Steuer AF, Storts DR, Thomson J, Taraporewala Z, Alston-Roberts C, Kerrigan L.  In Vitro Cell Dev Biol Anim. 2010 Jun; 10(6): 441-8.

Cell line cross-contamination initiative: an interactive reference database of STR profiles covering common cancer cell lines. Dirks WG, MacLeod RA, Nakamura Y, Kohara A, Reid Y, Milch H, Drexler HG, Mizusawa H. Int J Cancer. 2010 Jan; 126(1); 303-4.

Check your cultures! A list of cross-contaminated or mis-identified cell Lines. Capes-Davis A, Theodosopoulos G, Atkin I, Drexler HG, Kohara A, MacLeod RA, Masters JR, Nakamura Y, Reid YA, Reddel RR, Freshney RI. Int J Cancer 2010 Jul; 127(1); 1-8 Review.

Glucose-6-phosphate dehydrogenase mosaicism: utilization as a cell marker in the study of leiomyomas. Linder D, Gartler SM., Science 1965 Oct 1; 150(3692); 67-9