Using short tandem repeat profiling to validate cell lines in biobanks

Abstract

Aim. To approve the COrDIS kit (Gordiz, Russia) for the authenticity of cell lines from the Bioresource Collection of the N.N. Blokhin National Medical Research Center of Oncology by the short tandem repeat (STR) profiling.Material and methods. The chosen method proved to be a reliable and reproducible option. With this approach, a number of polymorphic STR loci are amplified using commercially available primer sets. Polymerase chain reaction (PCR) products are analyzed simultaneously with size standards using automated fluorescent detection methods. The results are presented as a simple number code corresponding to the lengths of the PCR products amplified at each locus. By applying this method to cell lines, the laboratory can both authenticate commercial cell lines and build a database of their lines. In the work, we used the COrDIS EXPERT 26 kit (Gordiz, Russia), validated for molecular genetic identification of personality based on multiplex PCR analysis of 26 highly polymorphic loci of human genomic deoxyribonucleic acid. PCR results were analyzed by capillary electrophoresis using an automatic genetic analyzer with laser-induced fluorescence detection (Applied Biosystems 3500xL).Results. When testing the method, profiling of 37 cell lines was carried out, of which 18 were announced in international databases and 19 were unique, obtained at the N. N. Blokhin National Medical Research Center of Oncology, as well as a cell line mixture in order to determine the limits of contamination detection. The obtained results showed the correspondence of commercial cell lines with the data in international databases. Within the framework of this work, profiles of unique lines were obtained and the foundation of own genetic database was laid. Studies to identify the limit of contamination detection by another line have shown that even 4% of the contaminant culture in the total pool can be used to identify its individual alleles.Conclusion. The results obtained indicate the possibility of using the method to identify samples of the collection and detect intraspecific contamination.

Human Metastatic Melanoma Cell Lines Panel for In Vitro and In Vivo Investigations

“…The amount of DNA was evaluated on a Qubit 4.0 fluorometer using the Qubit™ dsDNA BR Assay Kit (Thermo Fisher, Waltham, MA, USA). The authenticity of the cell lines (compared to the primary material) was confirmed by profiling based on short tandem repeats (STR) for 26 highly polymorphic loci using the COrDIS "EXPERT 26" kit (Gordiz, Moscow, Russia) [11]. PCR products were separated via capillary electrophoresis on a genetic analyzer with laser-induced fluorescent detection (Applied Biosystems 3500×L, Waltham, MA, USA) using 3500×L Genetic Analyzer 24-Capillary Array (Applied Biosystems, Waltham, MA, USA) of 50 cm length and POP-7 polymer (Applied Biosystems, Waltham, MA, USA).…”