Monitoring cell line identity in collections of human induced pluripotent stem cells

Abstract

The ability to reprogram somatic cells into induced pluripotent stem cells (hiPSCs) has led to the generation of large collections of cell lines from thousands of individuals with specific phenotypes, many of which will be shared among different research groups as invaluable tools for biomedical research. As hiPSC-based research involves extensive culture of many cell lines, the issue periodic cell line identification is particularly important to ensure that cell line identity remains accurate. Here we analyzed the different commercially available genotyping methods considering ease of in-house genotyping, cost and informativeness, and applied one of them in our workflow for hiPSC generation. We show that the chosen STR method was able to establish a unique DNA profile for each of the 35 individuals/hiPSC lines at the examined sites, as well as identify two discrepancies resulting from inadvertently exchanged samples. Our results highlight the importance of hiPSC line genotyping by an in-house method that allows periodic cell line identification and demonstrate that STR is a useful approach to supplement less frequent karyotyping and epigenetic evaluations.

Academic application of Good Cell Culture Practice for induced pluripotent stem cells

“…Therefore, leading cell banks (ATCC, CellBank Australia, DSMZ, ECACC, JCRB, and RIKEN) introduced the technique of STR profiling to address this issue (Pamies et al, 2017). According to the International Cell Line Authentication Committee (ICLAC), the analysis of at least eight STR loci is required for cell line authentication (Sarafian et al, 2018), while ISCBI recommends the use of the core 13 loci commonly used in forensic medicine (Xu et al, 2013). Commercially available kits on the market typically use a common subset of 16 different STR loci, which ensures comparison between different providers (Andrews et al, 2015).…”

“…Another approach is the analysis of single-nucleotide polymorphisms (SNPs), however they are discussed to be too detailed and expensive to be used for cell line authentication on a regular basis (Ntai et al, 2017). Comparing these two methods, individual STRs are more polymorphic (Freedman et al, 2015;Sarafian et al, 2018) and are widely applied in forensic analysis (Almeida et al, 2016), but spontaneous mutations or epigenetic changes due to long term culture (Lorsch et al, 2014) and the possible cross-contamination with cell lines from other species (e.g., mice) will not be detected (Freedman et al, 2015). 52-plex SNP assays seem to have the same rate of discrimination as 16-plex STR assays, but a centralized, online reference database for SNP assays is lacking (Freedman et al, 2015;Pamies et al, 2017).…”

Induced Pluripotent Stem Cells Provide an Unlimited T-cell Source for CAR-T cell Development and A Potential Source for Off-the-shelf Products

“…Giemsa-banding (G-banding) is a technique that produces a visible karyotype and used for chromosome counts to detect aneuploidies and karyotypically abnormal hiPSCs [70]. Besides, the fingerprinting technique analyses short tandem repeats (STR) loci to authenticate the hiPSCs and help to recognize unwanted switch or cross-contamination [71].…”

Clinical-Grade Human Pluripotent Stem Cells for Cell Therapy: Characterization Strategy

“…Identification of the particular hPSC line has to be unambiguous, therefore short tandem repeats (STR) loci are analyzed. There are numerous commercially available kits for cell line authentication [41,48], although for a few STR profiles establishment is worth consideration to send DNA samples to an accredited laboratory [37].…”