Platelet counting by the RBC/platelet ratio method. A reference method.

The International Council for Standardization in Haematology (ICSH) and the International Society of Laboratory Hematology (ISLH) recommend the counting of specifically labeled platelets relative to the RBCs with a fluorescence flow cytometer, together with an accurate RBC count determined with a semiautomated, single-channel aperture-impedance counter as a reference method for the enumeration of platelets. Fresh EDTA-anticoagulated venous blood specimens are measured within 4 hours of the draw. The specimen is prediluted (1:20) and the platelets labeled with two monoclonal antibodies specific to a cluster of differentiation common to all platelets. A final 1:1,000 dilution is made and at least 50,000 events with a minimum of 1,000 platelet events are counted with a flow cytometer to determine the RBC/platelet ratio. The platelet count is then calculated from this ratio and the RBC concentration of the original blood specimen.