title: Miniaturized flow cytometry-based in vitro primary human lymphocyte micronucleus assay-validation study.
abstract: Most in vitro mammalian genotoxicity assays show a low specificity (high rate of irrelevant positive results), and therefore, lead to an increase in follow-up in vivo genotoxicity testing. One of the sources of the high rate of in vitro irrelevant positive results that find no confirmation in in vivo studies may be the characteristics of the test system used. It has been shown that cells that are p53 deficient or carry an alteration in DNA repair genes may be more prone to produce high rate of false/irrelevant positive results. Primary human lymphocytes (HuLy) are considered to show a higher specificity in predicting the in vivo genotoxic potential of a tested compound. We recently developed a flow cytometry-based primary human T-lymphocyte micronucleus test (MNT) and showed that the technology is promising and reliable in detecting genotoxic compounds. The purpose of the present work was to develop and validate a miniaturized format of the assay. For validation purposes of the flow cytometry HuLy MNT a wide selection of compounds with different mechanisms of genotoxicity was used. The evaluation covered 30 compounds: 19 commercially available genotoxicants and nongenotoxicants and 11 early pharmaceutical development compounds. Being faster and less tedious than the microscopic analysis, the miniaturized flow cytometry-based methodology showed very promising results. Conveniently, cell division is verified within the same sample as the MN frequency. Moreover analysis of hypodiploid events may provide an indication for a mode of action, i.e. clastogenic versus aneugenic mechanism, for further follow-up testing.
authors.affiliation: Genetic toxicology and Safety Pharmacology, Preclinical Safety, Novartis Institutes for Biomedical Research, Basel, Switzerland.
authors.forename: Magdalena
authors.initials: M
authors.lastname: Lukamowicz-Rajska
authors.forename: Micheline
authors.initials: M
authors.lastname: Kirsch-Volders
authors.forename: Willi
authors.initials: W
authors.lastname: Suter
authors.forename: Hansjoerg
authors.initials: H
authors.lastname: Martus
authors.forename: Azeddine
authors.initials: A
authors.lastname: Elhajouji
mesh_terms: D000328:Adult; D002455:Cell Division; D005434:Flow Cytometry / Q000379:methods*; D006801:Humans; D066298:In Vitro Techniques; D015162:Micronucleus Tests / Q000379:methods*; D009153:Mutagens / Q000633:toxicity; D011003:Ploidies; D013601:T-Lymphocytes / Q000187:drug effects* / Q000473:pathology*
chemical_list: D009153:Mutagens