Comparative scoring of micronucleated reticulocytes in rat peripheral blood by flow cytometry and microscopy

Toxicol Sci. 2003 Aug;74(2):309-14. doi: 10.1093/toxsci/kfg143. Epub 2003 May 28.

Abstract

A flow cytometric technique for scoring the incidence of micronucleated reticulocytes in rat peripheral blood was compared to a standard microscopy-based procedure. For these studies, groups of five male Sprague-Dawley rats were treated with vehicle or a broad range of chemical genotoxicants: 6-thioguanine, N-methyl-N'-nitro-N-nitrosoguanidine, vincristine, methylaziridine, acetaldehyde, methyl methanesulfonate, benzene, monocrotaline, and azathioprine. Animals were treated once a day for up to 2 days, and peripheral blood was collected between 24 and 48 h after the final administration. These samples were processed for flow cytometric scoring and microscopy-based analysis using supravital acridine orange staining, and the percentage of reticulocytes and micronucleated reticulocytes was determined for each sample. The resulting data demonstrate good agreement between these scoring methodologies, although careful execution of the flow cytometric method was found to enhance the micronucleus assay by reducing both scoring time and scoring error. These data add further support to the premise that the peripheral blood compartment of rats can be used effectively to detect genotoxicant-induced micronuclei.

Publication types

  • Comparative Study
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Acridine Orange / metabolism
  • Animals
  • Antigens, CD / analysis
  • Antigens, CD / metabolism
  • Antigens, Differentiation, B-Lymphocyte / analysis
  • Antigens, Differentiation, B-Lymphocyte / metabolism
  • Cell Count / methods
  • Flow Cytometry / methods*
  • Fluorescent Dyes / metabolism
  • Image Cytometry / methods*
  • Male
  • Micronuclei, Chromosome-Defective / classification
  • Micronuclei, Chromosome-Defective / drug effects
  • Micronuclei, Chromosome-Defective / ultrastructure*
  • Micronucleus Tests / methods*
  • Mutagens / toxicity
  • Rats
  • Rats, Sprague-Dawley
  • Receptors, Transferrin
  • Reproducibility of Results
  • Reticulocytes / drug effects
  • Reticulocytes / metabolism
  • Reticulocytes / pathology*

Substances

  • Antigens, CD
  • Antigens, Differentiation, B-Lymphocyte
  • CD71 antigen
  • Fluorescent Dyes
  • Mutagens
  • Receptors, Transferrin
  • Acridine Orange