AN AUTOMATED-METHOD FOR DISCRIMINATING ANEUGEN-INDUCED VS. CLASTOGEN-INDUCED MICRONUCLEI

A Flow cytometric (FCM) procedure for quantitating micronucleated reti culocytes in mouse peripheral blood samples was evaluated for its abil ity to discriminate between aneugen- and clastogen-induced micronuclei (MN). In this experiment, BALB/c mice were infected with 0.9% saline, the model clastogen methyl methanesulfonate (100 mg/kg bw) or the ane ugen vincristine (0.2 mg/kg bw). Peripheral blood samples were collect ed 48 hr after infection and were subsequently fixed and stained For f low cytometric analysis. The staining method utilized FITC-conjugated anti-CD71 to differentially label reticulocytes, and the nucleic acid dye propidium iodide to resolve erythrocyte populations with and witho ut micronuclei. The frequency of micronucleated reticulocytes was dete rmined by analyzing 10,000 total reticulocytes per blood sample. A sec ond analysis was performed on each sample whereby the propidium iodide associated fluorescent signals of 250 MN were collected and graphed a s a single-parameter histogram. The histogram statistic ''median chann el'' was recorded For each sample and provided a quantitative descript ion of MN distribution according to DNA content. Cumulatively, the res ults of this study suggest that 1) flow cytometry con be employed to m easure the incidence of MN resulting from clastogenic or aneugenic act ivity, and 2) MN resulting from aneugens can be discriminated From tho se arising spontaneously or From clastogen treatment based on flow cyt ometric analysis of DNA content. (C) 1998 Wiley-Liss, Inc.