Question 7 of 20 Fundamentals Concept
Why must a cell line be authenticated, and how is it done?
Short answer
Misidentified and cross-contaminated cell lines have produced large amounts of published work about the wrong cell type. Human lines are authenticated by short tandem repeat profiling, which reads a set of highly variable DNA markers and compares the pattern with a reference profile.
Why
Short tandem repeats are short sequences repeated a variable number of times, and the combination across several markers is close to unique for a person. Comparing a culture's profile with the depositor's reference profile shows whether the cells are still what the label says. ATCC's guidance notes that without periodic testing, over-subcultured, misidentified or cross-contaminated lines reach the literature and produce spurious data, and recommends reporting the designation, catalog number and passage numbers in the methods.
Testing is done when a line arrives, when a new frozen stock is made, before a major experiment, and before publication. Journals and funders increasingly require the profile. The method has limits.
It identifies the donor, not the cell type, so two lines from the same patient cannot be told apart, and it cannot detect contamination of a human line by a mouse line, which needs an interspecies test. Profiles can also drift as a line loses or gains markers, so matching is scored as a percentage rather than as exact identity. Authentication also does not detect mycoplasma, which is a separate test.
A strong answer covers
- Misidentification and cross-contamination are common and consequential
- STR profiling compares variable repeat markers with a reference
- Test on arrival, at new stocks and before publication
- Report designation, source and passage number
- STR identifies the donor, not the cell type, and misses mycoplasma
Common mistakes
- Assuming a line bought from a repository stays authentic through years of passaging
- Treating an STR match as proof the culture is also free of mycoplasma
- Expecting STR to distinguish two lines derived from the same donor
The interviewer might ask next
What would you do if a profile came back as a partial match to the reference?