Cell Line Characterization

As a drug developer, the first manufacturing regulatory challenge faced is generation of a Master and Working Cell bank. Production must be carried out under Good Manufacturing Practice and requires specialist knowledge both for the manufacturing itself and the extensive biosafety and characterization testing required to release the banks.

Sartorius offers a broad range of services to characterize your mammalian cell banks in accordance with ICH regulatory guidelines (ICH Topic Q 5 D Quality of Biotechnological Products: Derivation and Characterisation of Cell Substrates Used for Production of Biotechnological/Biological Products). Our services comply with global pharmacopeial standards and are performed in facilities accredited by the U.S. FDA as well as the EMA and MHRA.

Ready-to-Use Quality Control Plans support testing and release of cell banks for use in manufacturing programs:

  • Research Cell Bank (RCB)
  • Master Cell Bank (MCB)
  • Working Cell Bank (WCB)
  • End of Production Cell Bank (EoPCB)  
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Integrated cGMP Biosafety & Characterization Services

Sartorius has extensive experience in characterization of cell banks used in production of biologic drugs for protein- and viral-based therapies. Sartorius’ QC testing plans meet requirements for mammalian cell bank release and are designed to meet ICH Q5A regulations with assays validated to GMP standards. Ready-to-use testing schemes may be added to existing Drug Development Risk Strategies or customized for the needs of specific programs.

Cell Bank Biosafety Testing and Characterization Packages

Cell Bank Testing Requirements

With a safety profile established over 20 years, CHO cells are by far the most used production cell lines in the pharmaceutical industry. Sartorius has performed biosafety and characterization testing on over 200 CHO cell banks. Our scientists have a wealth of experience to advise on the most appropriate and cost-effective testing strategy to meet regulatory requirements.  This recommendation takes into consideration both the history of the cell line and the raw materials used during the manufacture of the bank. Below is a standard QC testing plan for CHO cell line characterization.  While this plan may be modified for cells of human and simian origin, the basic principles apply.
 

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Test

MCB

WCB

EoPC

Identity

 

 

 

Mycoplasma

 

 

 

Sterility

 

 

 

In vitro assays

 

 

 

In vivo assays

 

 

 

Antibody tests

 

 

 

Retrovirus infectivity

 

 

 

Reverse transcriptase

 

 

 

Electron microscopy

 

 

 

Species specific tests

 

 

 

Table 1: Overview of GMP, GLP Testing Requirement

The process of cell banking begins with the introduction of a Research Cell Bank (RCB) into the manufacture of a Master Cell Bank (MCB).  

For this to occur, the RCB must be certified at a minimum to be sterile, mycoplasma free and identity assured. Additionally, it may be requested by the manufacturer that a broad-spectrum cell-based in vitro test for potential viral contaminants be performed.  

To expedite RCB release, options include rapid testing by Polymerase Chain Reaction (PCR), interim read in vitro strategies, and identity testing by DNA Barcoding.
 

Our team has extensive experience in the characterization of cell banks,  and provides a comprehensive set of assays for MCB testing. We can consult on the specific testing  requirements of a cell line or cell banks.  

Biosafety testing of the master cell bank encompasses a range of activities, including identity testing and evaluation of genetic stability. Additionally, a number of assays should be performed designed to detect the presence of potential contaminants.

A range of tests are used in the detection of contaminating agents, including those to detect microorganisms (sterility, mycoplasma and mycobacterium), as well as in vivo, in vitro assays, retrovirus infectivity assays, and transmission electron microscopy. These may be used to detect adventitious and endogenous viruses.

A series of species-specific virus tests is also required for the different production cell lines (Bovine, Porcine, Insect, Mouse, Hamster, Human, or Primate). To assess and determine the genetic stability of the cell bank, a number of different techniques can be applied. These may include Southern blotting, mRNA sequencing, or transgene copy number. Identity is determined by DNA barcoding.

Test methodology used to characterize master cell banks originating from different species include:

  • Sterility Testing
  • Mycoplasma Testing
  • in vivo assays
  • in vitro assays
  • Retrovirus assays
  • Bovine and Porcine assays
  • Electron Microscopy
  • Species-Specific assays


Genetic stability assays provided by Sartorius include the following:

  • Nucleic Acid Sequencing
  • Southern Blot
  • Identity Testing
  • Gene Copy Number

Contact Sartorius biosafety scientists to discuss MCB characterization requirements.

Working cell banks (WCB) are prepared from  the MCB. Biosafety testing requirements of the WCB depend upon the level of testing that has been conducted on the MCB. 

Where extensive characterization has already been completed on the MCB, minimal testing is required on the WCB. Depending on the regulatory authority, this is typically limited to assessment of identity, stability and contamination by microbes. Virus testing is only recommended where a recognized risk occurs. 

To characterize genetic stability, the transgene is assessed using techniques such as Southern blotting, sequencing of mRNA and/or transgene copy number. Identity can be measured using DNA barcoding. The presence of microbes is determined using sterility, mycoplasma and mycobacterium assays.

Test methodologies used to characterize working cell banks originating from different species include:

  • Sterility Testing
  • Mycoplasma Testing
  • in vitro assays

Genetic stability assays may be requested using these methods:

  • Nucleic Acid Sequencing
  • Southern Blot
  • Identity Testing
  • Gene Copy Number

An End of Production Cell Bank (EoPCB) is sometimes referred to as a Post-Production Cell Bank (PPCB) or as cells at the limit of in vitro cultivation. These cells are tested to validate the production system. This ensures that the cells are stable and that there are no contamination issues within the system.

The timing of the sampling for an EoPCB will vary between products and is performed on a case-by-case basis.

For cell lines used in the manufacture of protein-based therapies, the testing requirements are two-fold; they include tests for stability and tests for contamination (Biosafety). Testing schemes for cell lines used in viral vaccines or viral vectors may not require the same genetic stability assessments depending on the MCB generation process.

Requirements for the EoPBC are very similar to that for the Master Cell Bank. The EoPCB requires Genetic Stability testing and direct comparison with the MCB. Tests used include genetic analysis of the transgene using techniques such as Southern blotting, mRNA sequencing or transgene copy number. A number of different methods are used to test for potential contaminants including those for microbes (Sterility and Mycoplasma), adventitious virus (in vitro and in vivo, Transmission Electron Microscopy) and specific virus tests, as determined by the process.

Test methodology used to characterize EoPBC originating from different species include:

  • Sterility Testing
  • Mycoplasma Testing
  • in vivo Testing
  • in vitro Testing
  • Retrovirus Testing
  • Bovine and Porcine Assays
  • Electron Microscopy
  • Species-Specific Assays

Genetic stability assays provided by Sartorius include the following:

  • Nucleic Acid Sequencing
  • Southern Blot
  • Identity Testing
  • Gene Copy Number

Related Assets

Brochure: Chinese Hamster Ovary (CHO) Cell Banks Safety

Read Brochure
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What is Mycoplasma?

Mycoplasmas are the smallest self-replicating organisms currently known to science.

Read Blog Article
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ICH Topic Q5A (R1)

Quality of Biotechnological products: Viral safety evaluation of Biotechnology products derived from cell lines of human or animal origin.

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