Rapid quantitative analysis of protein tyrosine residue phosphorylation in defined cell populations in whole blood and bone marrow aspirates

We developed a rapid method for quantification of tyrosine phosphorylation in immunophenotypically defined cell populations in specimens of whole blood and unprocessed bone marrow. Samples were formaldehyde-fixed and cells were permeabilized. Phosphotyrosine residues and surface antigens were simultaneously stained by monoclonal antibodies and visualized by flow cytometry. The accuracy of the method was confirmed by demonstration of an increase of phosphotyrosine levels in pp60 ^v-src transformed fibroblasts. In blood of healthy donors, monocytes and granulocytes showed higher levels of phosphotyrosine than lymphocytes. CD34⁺ peripheral blood stem cells showed slightly increased tyrosine phosphorylation compared to autologous lymphocytes. Significantly elevated levels of phosphotyrosine were demonstrated in leukaemic blasts compared to lymphocytes ( P  = 0.01).