Introduction: Continuous cell lines are well-recognized models for the study of medical conditions, particularly for cancer. However, there are cautions to be aware of when using continuous cell lines, including the possibility of contamination which has far-reaching consequences on the results of the research. This is the reason why there are a lot of tests designed to help authentication of the cell line, which actually means finding out whether a certain cell line is “free“ from another cell line and/or microorganism or not. These kinds of tests are called authentication tests. The most frequently used one, also called “gold standard”, is a short tandem repeat profiling (STR) test. Objective: The aim of this study was to authenticate 6 cell lines grown in vitro using short tandem repeat (STR) profiling, as well as to define match percentage between electropherograms and ATCC- database. Materials and methods: 6 cell lines were used in this research (HeLa, SW620, CCRF-CEM, K-562, HT-29, Caco-2) and were analyzed on 15 loci + amelogenin. The steps of the analysis were: DNA isolation, determination of DNA concentration, PCR duplication and electrophoresis. GeneMapper software for electropherograms was used for the analysis of the results. Results: Electropherograms obtained were compared to standard reference STR database which made it possible to determine if cell lines were authentic and the percentage match between them. 4 cell lines, HeLa, SW620, CCRF-CEM and HT-29, were verified as authentic at 100 %, CaCo-2 cell line authentic at 75 %, while K-562 STR profile was not determined. Conclusion: Short tandem repeat profiling (STR) test established the authenticity for HeLa, SW620, HT-29 and CCRF-CEM cell lines, while further analysis should be conducted with Caco-2 cell line in order to make a final conclusion about its authenticity. Furthermore, analytical procedure for K-562 cell line should be repeated.