Development of a PCR assay to detect mycoplasma contamination in cord blood hematopoietic stem cells

Abstract

Background and Objectives: Contamination of cell lines and biological products is one of the major problems of cell culture techniques. Rapid detection of mycoplasma contamination in cell culture is an important part of quality control standards in related laboratories. The aim of this study was to evaluate the efficacy of PCR in detection of myroplasma as contaminants in cell cultures and other biological products. Method: PCR assays were optimized for 16 S rRNA target gene. Also the utilized PCR method was evaluated in terms of sensitivity and specificity. Finally, a simple DNA extraction and PCR analysis of 164 cell culture of adipose tissue derived mesenchymal stem cells were performed. Results: A 715 bp product was amplified and subsequently was confirmed by sequencing. The technique could detect 10 copies of the target DNA. No cross-reactivity with genomic DNA of other microorganisms was observed. Conclusions: The PCR technique in this study was based on 16S rRNA gene. It was highly sensitive and specific since it was able to detected Mycoplasma contamination in cell cultures

Keywords

• Mycoplasma
• PCR
• cell culture
• contamination
• molecular detection