FLOW CYTOMETRIC ANALYSIS OF BOVINE CD4 AND CDS LYMPHOCYTES - INFLUENCE OF BLOOD-SAMPLING AND PROCESSING METHODS

Technical information to facilitate bovine blood treatment for optimum lymphocyte flow cytometry analysis is reported. Murine monoclonal ant ibodies CC8 and CC63 were used to identify phenotypes corresponding to bovine CD4 T cells and CD8 T cells. Blood samples collected in acid c itrate dextrose (ACD) enhanced leucocyte subpopulation separation comp ared with ethylenediamine tetra-acetic acid, heparin and sodium citrat e. To preserve bovine blood before immunophenotyping, samples collecte d in ACD may be kept at 22 degrees C or at 4 degrees C and should be a nalysed within 32 hours. For isolation of white blood cells, whole blo od lysis was faster and gave the same results as Ficoll gradient separ ation 1.077 and Ficoll gradient separation 1.083. After immunophenotyp ing, blood could be stored at 4 degrees C if fixed with paraformaldehy de within seven days. Owing to diurnal variations, blood should be col lected at a standard time of the day.