What Tests Should You Include for Cell Line Characterization?

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The proper cell line characterization provides the groundwork for successful production of your biological product. These efforts are essential for product safety and quality because they provide documented evidence that your cell banks are free from contaminants and stable enough to allow you to continue your production processes years down the road.

Cell line characterization is performed for Master Cell Banks (MCB), Working Cell Banks (WCB), and End of Production Cells (EPC) or cells at the limit (CAL) of in vitro cell age used for production. According to current regulations, the following aspects must be considered:

  • The origin and history of the cell line Cellular morphology and growth characteristics
  • Cellular identity
  • Purity of the cell lines (i.e., absence of contaminating cells, microbial contaminations, and contaminations by adventitious viruses)
  • Tumorigenicity and oncogenicity
  • Stability

The regulatory requirements for the characterization of cell lines may vary between the different jurisdictions. Furthermore, testing recommendations depend on the species and origin of the cell line (e.g., bacterial, yeast, plant, insect, avian, or mammalian) as well as on the type of the product and its intended use (e.g., recombinant protein, vaccine, gene therapy product). When executing the assays, the guidelines recommend using state-of-the-art testing procedures which have been validated to show their suitability for the intended purpose. With all of these things to consider, it is important to determine not only the most effective but also the most relevant and efficient cell line characterization plan for your cell line. 

Charles River’s experts will help you to determine what characterization assays are specifically required for your product. We also recommend the consultation of national regulating authorities to ensure regulatory acceptance of the test program, especially for new cell substrates.

Cell Bank Timeline Planner

Gantt chart of cell bank production and characterization programThis interactive tool gives you insight into estimated timelines required for production and characterization, allowing you to plan your cell banking program timelines. Map Out Your Cell Bank

Cell Line Characterization Panels – Recommended Testing Plans

Validated methods for mammalian and microbial cell line characterization are available at Charles River sites in both the United States and Europe. References to guidelines concerning cell line characterization can be found in our regulatory library. The tables below list recommended testing plans for the characterization of general mammalian and microbial cell lines used to produce a recombinant protein along with a specific example for CHO cells.

Mammalian Cell Line Characterization Panel1 MCB WCB EPC/CAL
Microbial Contamination Sterility X X X
Mycoplasma X X X
Mycobacterium X    
Spiroplasma X    
Cell Line Identity DNA barcoding X X X
DNA fingerprinting/STR X   X
Karyology X   X
Genetic Stability DNA sequencing X   Optional
Gene copy number by qPCR X   X
Virus Testing Retroviruses: Reverse transcriptase and retroviral infectivity assays X   X
In vitro adventitious viruses X X X
In vivo adventitious viruses X   X

Test for specific viruses such as:

  • Bovine viruses
  • Porcine viruses
  • Human viruses
  • Mouse, hamster, and rat antibody production assays (MAP, RAP, and HAP)
X    
Transmission electron microscopy X   X
Tumorigenicity/Oncogenicity     X

1Novel molecular methods based on next-generation sequencing technology can also be offered to supplement the test program.

 

Microbial Cell Line Characterization Panel for MCB and WCB
Bacteriophage detection
Purity
Identification
Viability
Copy number
DNA sequencing
Retention of selectable markers
Retention of recombinant construct
Restriction endonuclease analysis

 

CHO Cell Line Characterization Panel MCB WCB EPC/CAL
Microbial Contamination Sterility X X X
Mycoplasma X X X
Cell Line Identity DNA barcoding X X X
Genetic Stability DNA sequencing X   X
Gene copy number X   X
Restrisction endonuclease analysis X   X
Virus Testing Endogenous and non-endogenous retroviruses: reverse transcriptase and retroviral infectivity assays X   X
In vitro adventitious viruses X   X
In vivo adventitious viruses (suckling and adult mice, embryonated eggs) X   X

Test for specific viruses such as:

  • Bovine viruses
  • Porcine viruses
X    
Mouse and hamster antibody production assays (MAP and HAP) X    
Transmission electron microscopy of cell thin sections X   X

For more information on our cell line characterization services or to schedule a consultation, please contact us at [email protected].