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Multicenter Evaluation of the 3% Paraformaldehyde Method for White Cell Counting in Leukocyte‐Reduced Red Blood Cells

Background/Aim: The 3% paraformaldehyde (PFA) method is a simple technique for counting residual white blood cells (WBC) in leukocyte‐depleted red blood cells (RBC). Preliminary data suggested that its sensitivity is at least equal to PCR and flow cytometry. We report the results of a multicenter study conducted by the BEST Working Party to determine precision and accuracy of the 3% PFA method. Study Design: In the 7 participating laboratories, 5 sets of samples containing nominal concentrations of 200, 100, 50, and 10 WBC/ml were prepared by diluting whole blood into ‘WBC‐free’ RBC. Ten milliliters of each sample were processed using the 3% PFA method, which is based on erythrocyte lysis and WBC concentration into 5% of the original sample volume; a Nageotte chamber is used to count concentrated WBC. Results: The precision of the technique varied according to the nominal concentration, ranging from a CV of 12% at 200 WBC/ml to 57% at 10 WBC/ml. The technique measured fewer than the nominal WBC concentrations (mean of all laboratories, ‐12.4%); underestimation was probably due to cell loss during sample manipulation. Overall accuracy was however acceptable, because statistical considerations establish that the actual WBC concentration would unlikely exceed 2 times the estimated count. Conclusions: The 3% PFA method is suitable for the enumeration of residual WBC at concentrations ≥50/ml. It represents a useful tool for evaluation of high performance filters by reference laboratories.