Mycoplasma contamination of manufacturing cell cultures is rare, but presents a threat for biologics manufacturers. Contamination sources can include raw materials used in the manufacturing of cell culture media, media, manufacturing personnel, and even the donors of cells used for cellular therapy products.

Recognition of the issue resulted in regulations to guide detection of mycoplasma in bioprocesses. Early guidelines stipulated application of mycoplasma tests based on lengthy culture, particularly the broth culture followed by plating on agar method.

Evolving Regulatory Guidance:

In 2007, the European Pharmacopoeia released guidance on performance expectations and validation of nucleic acid-based mycoplasma detection methods. The guidance on sensitivity is 10 colony forming units (CFU) or genome copies (GC)/mL of test sample. Since then, that has been the limit of detection (LOD) target that is generally applied when the goal is to replace the use of the 28-day test. The expectation on specificity is that a test for mycoplasma should not detect non-mycoplasma species.

Case Studies:

MycoSEQ Assay in the Real World

The largest privately owned plasma fractionator in the world, Octapharma manufactures medicines in the form of human Proteins sourced from human plasma and human cell lines.

• Many processes that used the MycoSEQ assay were being approved
• Thermo Fisher Scientific offered comprehensive support – training, instrument qualification services, assay optimization support, design of validation studies, and support for regulatory submissions.

Key Features

Sensitive

• Lowest limit of detection: 1–3 GC/qPCR reaction.
• Protocols designed to reduce background and concentrate mycoplasma from range of samples: raw materials, media, serum, cells, tissues, and large volumes.

Rapid

• Total test time is 5–6 h; standard method requires 28 days. Thus, the MycoSEQ assay accelerates production timelines for manufacturers of advanced biologics.

Cost saving

• The MycoSEQ assay costs ~$300/test; outsourced testing by the standard method costs ~$4000/test. Thus the ycoSEQ assay costs less than methods of lower sensitivity, specificity and speed.