A cell line is a population of cells cultured in a laboratory that can proliferate indefinitely, providing an accessible tool for research like cancer studies and drug development. For these studies to be valid, researchers must be certain they are working with the correct cells. Short Tandem Repeat (STR) profiling is a method that provides a unique genetic fingerprint to verify a human cell line’s identity, a process known as authentication.
The Critical Need for Cell Line Authentication
Cell line misidentification and cross-contamination are widespread problems in research. A substantial percentage of cell lines are not what researchers believe them to be, which compromises findings, leads to retracted publications, and wastes funding.
A well-documented example is the HeLa cell line, derived from Henrietta Lacks in 1951. HeLa cells are robust and have been used in biomedical research for decades, but their aggressive growth has also made them a common contaminant. Over the years, HeLa has overgrown and replaced hundreds of other cell lines, often without researchers’ knowledge.
This means a scientist believing they were studying breast cancer may have actually been studying cervical cancer cells from a HeLa contamination. This invalidates the specific study and can misdirect entire fields of research as other scientists build upon the erroneous data. Authentication serves as a safeguard against these costly errors.
The STR Profiling Mechanism
Short Tandem Repeats (STRs) are short, repeated sequences of DNA found at specific locations, or loci, in the genome. The number of repeats at any given locus is highly variable among individuals, creating a unique genetic signature used for identification.
Generating an STR profile begins with extracting DNA from the cell line and then amplifying the specific STR loci for identification. This is accomplished using polymerase chain reaction (PCR), which makes millions of copies of the STR regions from a small amount of DNA. For human cell line authentication, a standardized panel of core STR loci is used to ensure results can be compared consistently across labs.
After the STR regions are amplified, the DNA fragments are separated and measured using capillary electrophoresis. During this procedure, the amplified DNA fragments, which are fluorescently labeled, are passed through a very thin capillary tube containing a gel-like substance. An electric current is applied, causing the negatively charged DNA to move through the gel.
Shorter DNA fragments travel through the gel more quickly than longer ones, effectively separating them by size. A laser at the end of the capillary detects the fluorescent tags on each fragment as it passes by. This information is sent to a computer, which translates the fragment sizes at each STR locus into a series of numbers representing the cell line’s genetic fingerprint.
Analyzing and Interpreting an STR Profile
Once an STR profile is generated, it is compared against a reference profile. These reference profiles are maintained in public databases managed by major cell repositories, such as the American Type Culture Collection (ATCC) or the German Collection of Microorganisms and Cell Cultures (DSMZ). These repositories house authenticated cell lines and provide their official STR profiles for comparison.
The comparison uses an algorithm that calculates the percentage match between the alleles in the sample and reference profiles. A high degree of similarity indicates the cell line is authentic. A match of 80% or greater is required to confirm identity, which allows for minor genetic changes, known as genetic drift, that can occur as cells are grown in the lab.
A low percentage match signifies that the cell line is misidentified. Another possible outcome is a mixed profile, which contains more than two alleles at multiple STR loci. This indicates cross-contamination, where two or more cell lines are present in the same culture.
Implementing a Routine Testing Schedule
To maintain research integrity, STR profiling should be an integrated part of standard laboratory procedure, not a one-time event. Establishing a routine testing schedule ensures a cell line’s identity is verified at important points during its use. This proactive approach prevents the propagation of errors and safeguards the validity of experimental data.
Authentication should be performed upon receiving or creating any new cell line. This initial test establishes a baseline identity for the cells before any work begins. It is also important to test cells before freezing a new stock, or cell bank, to confirm the cells being preserved are correct.
Further testing is recommended if cells behave unexpectedly, such as showing changes in growth rate or appearance. It is also good practice to authenticate cells before starting a new series of experiments and again before submitting a manuscript for publication. Many scientific journals and funding agencies now require proof of cell line authentication, making it a prerequisite for publishing and securing grants.
Regular testing is a component of rigorous and reproducible science.