Mouse Lymphoma Thymidine Kinase (TK) Gene Mutation Assay

The mouse lymphoma assay (MLA) is used to detect gene mutations in L5178Y TK+/- -3.7.2C cells.

Screening, Regulatory and Mutagenic Assessment

  • Cultures are incubated with several concentrations of the test compound for three to four hours in the presence and absence of metabolic activation (S9) and for 24 hours in the absence of S9.
  • Cells are subcultured for two days following the end of treatment to allow expression of the mutant phenotype, and then seeded into 96-well plates in media with and without trifluorothymidine (TFT) to select for mutants and determine cloning efficiency.
  • Mutant colonies are characterized by size, since small and large colonies can be indicative of chromosomal events and gene mutations.
  • A positive outcome is characterized by a dose-dependent increase in mutant frequency that exceeds the Global Evaluation Factor (GEF).
Large and small colonies in an mouse lymphoma assay (MLA) plate
Large and small colonies in an MLA plate

When to Perform the Mouse Lymphoma Assay

  • Screening
  • IND-enabling
    • As part of the ICH S2(R1) standard battery
  • REACH requirement
    • As part of Annex VIII testing
  • Other uses
    • To assess mutagenicity when the Ames assay may not be appropriate (e.g., antibiotics, nanomaterials)