The Challenges of Mycoplasma Testing for USP <63> Compliance

by | Pharmaceutical

Publish Date: October 23, 2020

As the use of cell lines to produce biopharmaceuticals continues to increase, Mycoplasma testing is a growing quality control challenge. Because of its unique properties, Mycoplasma poses many challenges for detection and management. Mycoplasma contamination can spread quickly, yet common quality control methods can be time-consuming and complicated. We developed Enumerated Mycoplasma to simplify compliance with USP <63>. In addition to unique controls, QC for Mycoplasma also requires different methodologies because of its colonization characteristics and other factors outlined below. We asked R&D Scientist Karla I Ziegelmann-Fjeld, PhD and Technical Support Manager, Kali Sorum, RM(NRCM) to address common issues surrounding USP <63> to help streamline your Mycoplasma QC.

USP <63> FAQ

Why do I need to validate the product with the media I plan on using?

Microbiologics conducted a study with three different medias and has collected feedback from customers. This information shows that the CFU count can vary significantly based on the media used for the experiment. 

The CoA for this product says that it is tested on SP4 Glucose Agar. If I also use SP4 Glucose Agar do I still need to validate the product with my media?

Yes. It will still be very important to do the recommended validation. SP4 Glucose Agar can vary from manufacturer to manufacturer and have different antibiotics or other additives that could potentially significantly impact the CFU count. Additionally, if the plates are grown in an atmosphere that differs from the atmosphere used at Microbiologics (i.e. lower oxygen concentration, cooler incubator temperature) those factors could affect the recovery and growth time.

How many plates can I inoculate from a single tube?

This will depend on the outcome of your validation. If your media shows that the mean CFU value is in the E04 range, then you will be able to dilute the product to inoculate many plates. If your media reduces the count to a E03 or E02 count then you will not be able to dilute the product as much, or perhaps at all, so you will be able to inoculate fewer plates.

Can I use this product as a control for tests/assays other than QC of plated media?

This product is a suspension of frozen, viable cells. It can potentially be used for multiple purposes such as molecular based assays.

Is it ok if I thaw and refreeze the product for later use?

Freeze-thaw cycles are very stressful to the organisms. Microbiologics cannot guarantee the enumeration of the product if it is thawed and refrozen.

How do I visualize the mycoplasma cells?

Mycoplasma colonies are much smaller than most other bacterial colonies. Some may be visible to the naked eye, but colony characteristics are only visible under a microscope.

What are the growth conditions for each strain?

Microbiologics follows the growth conditions of USP <63> internally and recommends them for customer use.


Use of Enumerated Mycoplasma Controls to Assess Media Quality for Growth Promotion Testing.

Enumerated Mycoplasma Media Difference Information.

What USP <63> Challenges Does Your Lab Face?

We value your input. Please take our short (5-7 minute) survey on Mycoplasma testing to help us continue developing the products you need for USP <63> compliance.

Written by Microbiologics

Microbiologics is the foremost biological products and services provider, focused on protecting the health and safety of people around the world. For over 50 years, they have been partnering with healthcare and life science laboratories, manufacturers and suppliers across the globe to co-create and provide biological control materials, assay services and consulting for microbiology, molecular diagnostics and virology. Headquartered in Saint Cloud, Minnesota, the company has additional facilities in California, Kentucky and Michigan. Microbiologics has advanced industry standards for biological controls by adopting the most rigorous testing and quality standards including ISO 13485, ISO/IEC 17025, and ISO 17034. The company has an extensive distribution network spanning over 150 countries. For more information, visit

You May Also Like


Share This