Mycoplasma, Sterility Testing and LAL

Mycoplasma Detection

Mycoplasma (mollicutes) grow in the presence of cells. According to the regulatory agencies cells need to be checked for contamination.

The FDA note “Points to Consider on the Characterization of Cell Lines Used to Produce Biologicals” (1993) advises detection of culturable and non-culturable Mycoplasma using indicator DNA fluorochrome staining and by culture.

Texcell has developed a protocol capable of detecting a broad range of Mycoplasma species encountered in cell lines of human and animal origins.

Under certain circumstances, the client will need to control pre-banking material or products for the presence of mycoplasma without specific recommendations. By request, Texcell can use an indicator cell line to grow specific Mycoplasma and then detect them by fluorescent staining of cultured cells after inoculation with a test product.

Detection of Mycoplasma by culture is time consuming, requiring at leats 28 days for cultivation of mollicutes. Texcell has developed a quantitative Polymerase Chain Reaction (qPCR) able to detect more than 90 species.

Sterility Evaluation

The US and EU Pharmacopoeia or FDA PTC guidelines describe several techniques for the detection of bacteria in cells or cell-derived products.

In accordance with the ICH Q5D, Texcell has developed specific assays for unprocessed bulks, cell banks and cells at an age suitable for in vitro production.

The purpose of sterility tests is to detect microbiological contaminants such as bacteria, yeast and fungi. Myclopasma detection requires special protocols (see section above “Mycoplasma detection”)

Limulus Amoebocyte Lysate Test

The LAL test for pyrogenicity involves the detection of gram-negative bacterial endotoxin by catalisying activation of the proenzyme. This test prevents the risk of ferbrile reaction, shock and death in patients following the injection of a test product.