Millipore Technical Publications |
 | Application Note - SDS1700ENEU | |
Mycoplasma Testing | ||
| Lit No: | SDS1700ENEU | |
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Mycoplasma is the generic name assigned to microorganisms in the class Mollicutes. It comprises over 100 known species that are further divided into a number of strains. Mycoplasmas are the smallest, free living, self-replicating microorganisms. Mycoplasma contamination is of greatest concern in the area of cell culture. It has been estimated that between 5 and 35% of cell cultures currently in use are infected with at least one species of mycoplasma1. The highly nutritive environment of the media used in cell culture can lead to the propagation of mycoplasma, resulting in diminished cell growth as well as the loss of cultures. Because they do not overgrow the cell lines, specific methods are necessary to detect them, often requiring a combination of tests.
What Products Should Be Tested?
Therapeutic products for both human and animal use are required to undergo mycoplasma testing throughout various steps in the production process including:
- Cell lines/banks
- Virus seeds
- Fermented samples
- Crude harvests
- Bulk vaccines
- Purified biological products for human use
When Should Testing be Performed?
Each lot of product harvest concentrate should be tested prior to clarification, filtration, purification, and inactivation, although this stage of the manufacturing process may not be appropriate for all products.
1Hay, R.J., Macy, M.L., and Chen, T.R. “Mycoplasma Infection of Cultured Cells”. Nature (London), 229: 487 – 488, 1989.
Study Design
Assay consists of a culture method and an indicator cell test. Validation of the method using 6 strains is performed as part of the assay. Assay controls are selected according to product type, manufacturing process and customers’ specific needs. Both test methods meet the regulatory requirements listed under regulatory compliance.
Culture Test on Agar
- Inoculation in broth and agar media broth
- Several readings and sub-passages
- Incubation at 2 conditions
Assay Duration: 28 – 35 days
Cell Culture Method Using Indicator Cell
- Inoculation on indicator cell lines
- Detection of mycoplasma in cell culture by DNA fluorescence staining
- Examination for extra-nuclear fluorescence for mycoplasma
Assay Duration: 7 – 10 days
Sample Requirement: Dependent on customer supplied product
Regulatory Compliance
- EP 2.6.7 European Pharmacopeia (2.6.7 Mycoplasmas)
- American guideline 21 CFR 610.30 Code of Federal Regulations 21 CFR Ch. 1 (4-4-01 edition) § 610.30 Subpart D-mycoplasma
- Center for Biologics Evaluation and Research, FDA, Points to Consider in the Characterization of Cell Lines Used to Produce Biologicals 1993.
Final Report
MicroSafe Services from Millipore can provide each client with the following final reports:
- GLP report that includes all raw data, protocol methods and final result
- GMP report that includes a certificate of results
Custom Assays
We understand that no production plant is identical and that every process is unique. Using a combination of practical experience and a scientific approach, we will customize mycoplasma testing methods to meet your specific requirements including scientifically sound data that will be accepted by regulatory authorities around the globe.
Your Partner in Microbiology
MicroSafe Services from Millipore delivers a complete range of critical testing and consultancy services for the development and production of biopharmaceuticals. Leaders in the field of microbiology including in vitro virology testing, our staff of accomplished scientists, engineers and microbiologists will help you meet your specific critical testing requirements throughout every phase of drug production. All testing is conducted under GMP and/or GLP certifications as well as our own internal quality system.