COMPARISON OF FLOW CYTOMETRIC AND MANUAL BONE-MARROW DIFFERENTIALS INWISTAR RATS

Preclinical drug trials frequently require the evaluation of animal bo ne marrow, a time-consuming process requiring the skills of a highly t rained hematologist. In the present study, a flow cytometric technique was developed that could effectively replace the need for manual bone marrow differentials in rats. Peroxidase activity, measured indirectl y with 2'7'-dichlorofluorescin, was coupled with the use of species-sp ecific T-and B-lymphocyte antibodies and cell size to produce a flow c ytometric analysis of rat bone marrow, Accurate identification of lymp hocyte, proliferating and maturing erythroid and myeloid, and megakary ocyte populations was confirmed by cell sorting. Flow cytometry yielde d differentials that were indistinguishable from manual differentials and published reference ranges. Enumeration of lymphocyte numbers with monoclonal markers is a key advantage of flow cytometric differential s because misidentification of lymphocytes in poorly prepared or stain ed bone marrow smears is a common problem. The most apparent advantage is increased throughput and reproducibility. Operator training for an alysis using flow cytometry can be readily accomplished within a few d ays as opposed to the extensive training required for individuals perf orming manual bone marrow differentials. This methodology provides a h igh-volume, rapid, and relatively low-cost tool for the reliable evalu ation of rat bone marrow differentials that has been heretofore unavai lable. (C) 1998 Wiley-Liss, Inc.