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Are you using authentic cell lines?

Cell lines are incredibly valuable tools for biomedical research. However, their usefulness as models of normal or diseased tissues relies on them being truly of the tissue type and species that researchers think they are. Unfortunately, an estimated 14–46% of most commonly used cell lines are misidentified [1]. Misidentification often comes from cross-contamination (a culture being taken over by another cell line that is also cultured in the same laboratory) and may come from errors in handling the cells. HeLa cells, the first human cancer cell line ever established, are known to be extremely resilient that they can survive in aerosol droplets and contaminate other cells that way. Since HeLa cells grow very fast, they may eventually overgrow the other cells. Studies have found 30–50% of misidentified cell lines to be HeLa derivatives [2, 3]. For most misidentified cell lines, authentic stocks of those lines can no longer be found.

Even though the issue of cell line misidentification has been recognized since the 1960s, this continues to be a widespread problem. For example, HEp-2 was shown to be a HeLa derivative (cervical cancer cells) in 1968, but papers are still published every year with researchers using it as a laryngeal cancer cell line [1]. For a long time, journals and funding organizations did not have policies in place that require scientists to authenticate the cell lines they use. Things have changed in the last decade, as more and more journals now require or encourage authors to perform authentication of cell lines [1, 4], including the BioMed Central journals, International Journal of Cancer, Nature Publishing Group journals, and many others. Because the list is growing and requirements differ between journals, it is important to check a specific journal’s guidelines before submission. For example, in addition to a general requirement of cell line authentication, the Journal of Hepatology also states that it will no longer consider experiments using some known misidentified cell lines (BEL7402, SMMC7721, MHCC97L, etc). The US National Institutes of Health is one of the funding agencies that now mandate cell line authentication as a condition for funding.

The method of human cell line authentication accepted by journals is short tandem repeat (STR) profiling, which is a DNA fingerprinting technique used in forensics. It is a PCR-based technique, where a number of polymorphic microsatellite loci are tested simultaneously. STR profiling can match a cell line to a donor (if the donor’s STR profile is known), or a cell line with a known STR profile. However, if more than two cell lines have been intentionally established from the same donor, the two cell lines are not distinguishable by STR profiling. Because of genetic drift as cells are passaged, it is possible for a STR profile of cultured cells to not match that of the donor 100%. Current standard is to use a match ≥ 80% to confirm the identity of a cell line [5].
For more information on this issue and the recommended practices of using cell lines, check out the website of the International Cell Line Authentication Committee (ICLAC). ICLAC hosts a register of currently known misidentified cell lines, which now lists 488 cell lines. You can also find links to databases of human cell line STR profiles and many other publications and websites on this issue.

Do you know what cells lines you have?

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