New flow cytometric method for surface phenotyping basophils from peripheral blood.

To clarify the role of basophils in the pathogenesis of allergic disease, we developed a new method for performing surface phenotyping of these cells in centrifugation-enriched mononuclear cell fraction. This method identified basophils on the basic of a negative reactivity with mixed FITC-conjugated monoclonal anti-bodies (mAbs) (anti-CD2, -CD14, -CD16, and -CD19) with analysis performed by flow cytometry. The validity of this approach was confirmed by sorting experiments. Various PE-conjugated mAbs were also used to examine binding to FITC-negative basophils. Basophils from asthmatic patients (n = 14) as well as from normal subjects (n = 6) were shown to express CDw32 (Fc gamma RII), CD25 (IL-2R), but not CD64 (Fc gamma RI). We also detected binding of IgG1 and IgG4 to basophils. This method of phenotyping was very rapid and simple. It thus appears to be useful in the study of allergic disease, as well as of the biology of the basophil.