Adventitious Agents / Virology Testing

Adventitious agents are a major concern when using mammalian cell lines for production of a biotechnology product or biological. BioReliance performs the classic in vitro assay designed to detect adventitious viruses in cell banks, virus seed stock, raw materials and final products, including:

  • Adventitious virus assay
  • Porcine and bovine adventitious virus assay as described in the 9 CFR
  • A range of assays addressing specific contamination risks also is available

The BioReliance CRO team is available to help you design and execute the testing services needed to ensure your biologicals are free of viral contamination. BioReliance experts also are available for consultation on development and validation services for emerging viruses.

To see our in vitro testing services for adventitious agents, use the search tool above or click here to contact us.


Details about our in vitro adventitious virus assays

The goal is to create the best possible chance of catching contamination in your production cells. Each in vitro assay is fully validated in accordance with ICH Q2 (R1) guidelines and EP 2.6.21.

In vitro assays can be configured for testing serum, cell lysates, enzymes, raw materials, and other non-serum samples for the presence of adventitious agents by inoculating indicator cell lines with the above samples. Cultures are then maintained for 21 days and monitored for cytopathic effects (CPE).

Cultures exhibiting CPE immediately are harvested, fixed and screened for viruses. After 21 days, all remaining cultures are harvested examined for the presence of specific viruses by fluorescent antibody staining techniques. Further, the cultures are assayed for hemadsorption (hemagglutinin on the surface of viruses adsorbs to red blood cells (RBCs), resulting in the clumping of RBCs) to detect viruses that are difficult to see by fluorescence.

Does BioReliance offer other virology testing assays for adventitious agents?

BioReliance offers a wide range of services to enable production of biologicals free of contamination including:

The BioReliance CRO team has a broad experience in testing and detecting adventitious agents using a wide range of methods. The BioReliance research staff is available to support your virology testing needs from design through execution.


Virology Testing (adventitious virus)
BioReliance performs a series of biosafety assays designed to detect species specific and adventitious viruses in working cell banks, master cell banks, virus seed stock, raw materials and final products.

Each assay method is fully validated in accordance with ICH Q2 (R1) guidelines and EP 2.6.21. Assays are available for human viruses, respiratory viruses, porcine viruses, simian viruses, canine, murine and many more. Development and validation services also are available for emerging viruses.

BioReliance CRO’s virology testing services provide reassurance that the production of biologicals is free of contamination through assays including

In vitro assays - Virology Testing

  • In vitro detection of endogenous and adventitious agents
  • In vitro assays for presence of bovine and porcine as described in the 9 CFR
  • Custom assays based on your virology testing needs

MAP, HAP, RAP

  • Mouse antibody production test (MAP)
  • Hamster antibody production test (HAP)
  • Rodent antibody production test (RAP)
  • Custom assays based on your virology testing needs

Retrovirus Testing

  • Co-cultivation (murine, ecotropic, amphotropic and rodent)
  • PCR/RT-PCR detection of endogenous and adventitious agents Direct and extended XC plaque assay
  • Direct and extended S+L- focus assay (mink, feline)
  • Amplification in Mus Dunni cells
  • Transmission Electron Microscopy (TEM)

To see our virology testing services, use the search tool.

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Why does virology testing need to be performed on biologicals?

When production of a biotechnology product or biological is derived from mammalian cell lines, contamination by endogenous and adventitious viruses can occur during the manufacturing process. For example, many cell lines are grown in the presence of fetal bovine serum raising the possibility of a native bovine virus contaminating the end product.

The most common assays are in vitro virology tests in which the agent is inoculated into susceptible cell lines such as Vero cells, and the readout is a visible cytopathic effect, often called CPE in the literature.

Not all viruses show CPEs, so we recommend secondary detection methods like those outlined above. Below we detail a few common secondary virology testing methods. To see our description of other assays please follow the links above or just use our assay search engine above and to the right.


Quantitative PCR Testing to Detect Adventitious Virus

BioReliance performs quantitative PCR tests designed to detect adventitious and endogenous viruses in cell banks, virus seed stock, raw materials and final products. Assays are available for:

  • human viruses
  • respiratory viruses
  • porcine viruses
  • simian viruses
  • canine viruses
  • murine and many more specific target PCRs.

The BioReliance has broad experience in virology testing and is available to provide guidance in protocol design for the most appropriate collection, preparation and evaluation of biologic samples to ensure sample integrity for PCR detection of adventitious virus.

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Retrovirus Testing Services from BioReliance

Adventitious viruses are often thought of as purely retroviruses. Retroviruses are unique in that they can integrate into the host genome making them difficult to detect by normal methods.

To address this, the BioReliance provides a wide range of retrovirus detection testing services including:

  • XC plaque assay
  • S+L- focus assay
  • PERT
  • PCR/RT-PCR
  • Transmission electron microscopy (TEM)
  • Co-cultivation assays (murine, ecotropic, amphotropic and rodent)

Consult with our experts to determine the best methods for testing your therapeutics for the presence of retroviruses.

Co-cultivation Assays for Adventitious Virus

Co-cultivation of production cells at the limit of the in vitro age is an assay we offer to test for latent retroviruses for our clients. Essentially, your production cells are plated along with susceptible cell lines that exhibit stronger phenotypic CPE when the virus enters them. Thus, viruses that are expressed as non-infectious particles or exhibit low level of expression in production cells are detected.

Contact us to consult about specific testing for retroviruses by co-cultivation assays.

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Mouse Antibody Production Test

BioReliance performs in vivo assays designed to detect adventitious and endogenous viruses in cell banks, virus seed stock, raw materials and final products.

A major concern when using mammalian cell lines for production of a biotechnology product or biological is the risk of contamination by endogenous and adventitious viruses, which can occur during the manufacturing process. Such contamination could have serious clinical consequences detrimental to the patient. BioReliance in vivo virus detection services are available to ensure the absence of human and animal adventitious viruses in your biologicals.

In vivo testing services for viral contamination includes:

  • Mouse antibody production (MAP) test
  • Hamster antibody production (HAP) test
  • Rat antibody production (RAP) test
  • S+L- focus assay

The BioReliance CRO team has a broad experience in testing and detecting adventitious agents using in vivo methods, and research staff is available to provide guidance in protocol design for the most appropriate collection, preparation and evaluation of biologic samples to ensure sample integrity.

To see our in vivo testing services for adventitious agents, use the search tool above.

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Electron Microscopy Services - Adventitious Agents

Electron microscopy services provide high-resolution images of viral or biological specimens at an ultrastructural level to detect the presence, absence, and quantitation of known microbial or adventitious agents in biological fluids or in cell substrates.

BioReliance offers a variety of assay options for your electron microscopy needs including:

  • Ultra-Thin Section Electron Microscopy
  • Virus Particle Counts
  • Virus Characterization
  • Negative Stain
  • Immuno-electron microscopy
  • Bioprocess Validation

We provide GMP-compliant transmission electron microscopy (TEM) services for ultrastructural pathology and viral particle identification, tabulation, quantification and characterization in support of the FDA Points to Consider (PTC) documents. We can provide a fast turn around time and the highest quality testing. Our experienced CRO research staff is available to provide guidance in study design for the most appropriate collection, preparation and evaluation of biologic samples to ensure sample integrity.

To see our electron microscopy services, use the search tool above or click here to contact us.

How are viral particles characterized by transmission electron microscopy (TEM)?

BioReliance examines biological fluids and quantifies virus particles to estimate the viral load of the sample. It is essential that viral load is determined prior to validating the ability of a purification process to inactivate or remove viral contaminants.

Morphology of virus particles, as defined by transmission electron microscopy, is one of the major criteria for classification of viruses. Quantitation of virus particles can be performed by thin section TEM or by negative stain TEM.

What is Ultra-thin Section Electron Microscopy?

Ultra-thin sectioning reveals fine details of cell and tissue ultrastructure as well as the structure of viruses or other present microbes. Cells or tissues embedded in plastic are thin sectioned, mounted and stained, and examined by transmission electron microscopy at an ultrastructural level.

This technique provides details of the interior of the cells and tissues, including organelle structure, morphology of macromolecules, such as virus particles, without causing distortion of the cells or tissues.

Immuno-electron microscopy and intensive search for new or suspected viruses and other microbes are available as ultra-thin section EM services.

To see our electron microscopy services, use the search tool above or click here to contact us.

When is negative staining performed?

Negative staining is useful for quantitative and qualitative studies of viruses or other microbes. Specimen preparations are stabilized in electron-dense salts and examined by transmission electron microscopy.

Viral particle counts (VPC), immuno-electron microscopy, are also available as negative stain EM services.

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