Establishment of 100GC Mycoplasma Standards Based on Nucleic Acid Amplification Technology According to the Newly Revised European Pharmacopoeia 2.6.7 and the USP <77>

Mycoplasma contamination is a concern in cell culture or cell culture media during biopharmaceutical manufacturing processes. These impurities pose a safety concern in Advanced Therapy Medicinal Products (ATMPs) due to their small size and resistance to common antibiotics used in cell culture. The minimum required detection limit for mycoplasma detection methods for pharmaceutical products is traditionally defined as 10 colony-forming units (CFU). This definition is based on the classical culture method for detecting and quantifying Mycoplasma. With the advent of novel methods for Mycoplasma detection, which are faster and more reliable, the classical culture method has been superseded by Nucleic Acid Amplification Technology (NAAT). Therefore, the new revised EP 2.6.7 and USP <77> allow mycoplasma detection based on NAAT. Considering this new NAAT-based approach, new validation standards are needed that are based on genomic copies rather than colony-forming units. 100GC mycoplasma standards have been developed according to the revised European Pharmacopoeia 2.6.7, which are quantified using a novel, validated digital PCR assay. This assay offers advantages in accuracy and sensitivity compared to classical real-time PCR.