Protocol recommended by the CSGMT/JEMS.MMS for the short-term mouse peripheral blood micronucleus test. The Collaborative Study Group for the Micronucleus Test (CSGMT) (CSGMT/JEMS.MMS, The Mammalian Mutagenesis Study Group of the Environmental Mutagen Society of Japan).

Abstract

Although the target cells for the bone marrow (BM) and peripheral blood (PB) micronucleus tests are the same, erythroblasts, the PB method offers important advantages over the BM method. We propose a protocol for the short-term peripheral blood micronucleus test. This assay is intended primarily for the identification of a wide variety of chemical clastogens and spindle poisons, and secondarily for risk assessment. The recommended experiment size, seemingly small, has adequate detection power. Experimental results obtained from Collaborative Study Group for the Micronucleus Test (CSGMT) studies and data collected from a survey of the literature provided the basis of the proposed protocol. Our protocol, designed for mice, includes the following features. (i) The maximum tolerated dose (MTD) is determined experimentally with a small number of animals treated i.p. or per os (or by other routes, if called for) in a dose-finding test, which can be conducted simultaneously with tests for finding both number of treatments and optimal sampling time. (ii) At least three groups of five mice (at least four effective animals per group), males or females, are given i.p. or per os doses of, for example, the MTD, 1/2 MTD and 1/4 MTD once, twice or more, 24 h apart. (iii) Peripheral blood samples are taken before treatment (the 0 time control) and twice at 48 and 72 h for a single treatment, once between 24 and 36 h after the second treatment for double treatments, or once 24 h after the final treatment for multiple dosing. Or, if an optimal sample time is established in a preliminary test, samples are taken at that time, (iv) Samples are stained with acridine orange, and 2000 immature erythrocytes per animal are examined. (v) The combined data of 0 time samples are the negative control.(ABSTRACT TRUNCATED AT 250 WORDS)