PHASE TWO OF AN INTERLABORATORY EVALUATION OF THE QUANTIFICATION OF RAT SPLENIC LYMPHOCYTE SUBTYPES USING IMMUNOFLUORESCENT STAINING AND FLOW CYTOMETRY

Abstract

In phase one of an interlaboratory study, baseline values for rat splenic lymphocyte populations were established. In phase two, rat splenic lymphocyte populations were evaluated using immunofluorescent staining and flow cytometry following exposure to the immunosuppressive agent cyclophosphamide (CY). The study involved four independent facilities employing a common protocol. All laboratories purchased animals and reagents from the same sources. The objective of phase two was to determine whether each laboratory could detect a significant change in the same splenic lymphocyte population(s) at the same or similar CY dose levels. Crl:CD R BR male rats were dosed by the intraperitoneal route with 1, 3, or 10 mg / kg CY for 4 days. On day 5, spleen cell number and weights were obtained and splenic lymphocytes were evaluated following the lysis of red blood cells with ammonium chloride. Splenic lymphocyte populations were enumerated with monoclonal antibodies using the dual labeling of T-cell subpopulations a...