Big Blue® Transgenic Rodent Mutation Assay

BioReliance’s Big Blue® Transgenic Rodent Mutation (TRM) assay is used to measure mutations in any in vivo tissue - including germ cells – and is performed in a novel line of transgenic animal models. The role of Big Blue® is to investigate test articles believed to be mutagens and resolve their mechanism of action. A positive Ames test and negative cytogenetic tests usually lead to a TRM assay.

Originally co-developed and validated by BioReliance in the 1990s with funding from the National Institute for Environmental Health Sciences (NIEHS), Big Blue is currently in demand due to recent changes in regulatory guidelines and acceptance. As such, the assay has been re-qualified for commercial use by BioReliance and is defined by the OECD Testing Guideline 488” Transgenic Rodent Somatic and Germ Cell Gene Mutation Assays.”

  • OECD Test Guideline 488 has been approved

  • Recommended by ECHA for REACH registration

  • Indicated by ICH S2(R1) as a follow-up to positive in vitro gene tox mutation assays

  • Recommended in draft ICH M7 as primary follow up assay for impurities with mutagenic mode of action

The Big Blue® Transgenic Rodent Mutation Assay (TRM) is an in vivo gene mutation assay available in mice and rats. Gene mutation assays provide unique insights into chemicals that might have mutagenic modes of genotoxic action because they specifically measure gene mutations. Other commonly used assays only measure DNA damage at the chromosome level; however, DNA damage does not always result in mutations.

Consult with one of our genotoxicity experts to determine the toxicology assays that are best for your product needs or use the search tool on the right to browse our Big Blue® offerings.


Big Blue® BasicsBig Blue Flow chart

Big Blue® mice are bred to have multiple copies of recoverable target genes integrated into their genome. The animals are created by a microinjection of the lambda shuttle vector containing the cII gene into the pronucleus of fertilized eggs from either C57BL/6 mice or Fisher 344 rats. The principle of the assay is that we can recover the lambda shuttle vector from genomic DNA and measure mutations in the lambda cII gene.

In a typical study, animals are dosed by oral gavage for 28 days. On Day 31, after a three day period allowing DNA lesions to be fixed into the DNA and expressed as stable mutations, tissues are collected. After collection, high molecular weight DNA is extracted from tissues of interest and purified, the shuttle vector recovered, packaged into phage, infected onto E. coli, and plated for plaque formation at two temperatures. cII mutant phage form plaques only at 24°C; all phages form plaques at 37°C. Mutation frequency is calculated by dividing the number of mutant plaques by the total number of plaques. Bioavailability (TK) measurements can be added to demonstrate exposure.

OECD Testing Guideline 488 allows for differing designs. Please consult a BioReliance representative to design a study that will best fit your needs according to regulatory guidelines.

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