Distribution of cell-associated prion protein in normal adult blood determined by flow cytometry

Leucocyte subpopulations from normally healthy individuals were identified by recognized combinations of fluorochrome-conjugated antibodies to CD mark ers and stained by different monoclonal antibodies (MAb) to normal cellular prion protein (PrPC, including the 3F4 MAb Cell preparations were examined by three-colour now cytometry. All mononuclear leucocyte subpopulations an d platelets expressed PrPC, but polymorphonuclear leucocytes and red blood cells expressed little or no PrPC. The amounts of PrPC expressed by the dif ferent cells were calculated by comparison to bead standards, Mononuclear l eucocytes expressed 3000-4000 molecules of antibody-reactive PrPC per cell. Resting platelets expressed around 1400 molecules of PrPC per cell, wherea s activated platelets expressed around 4800 molecules of PrPC per cell. Ext rapolation of these values to the amounts of the various cells in whole blo od showed that platelet PrPC accounted for at least 96% of cell-expressed P rPC in blood. The PrPC on mononuclear cells and platelets was sensitive to enzymatic treatment of cells by proteinase k and phosphatidylinositol-speci fic phospholipase C. Certain anti-PrPC MAbs which showed equivalent intensi ty of staining to MAb 3F4 on fresh cells showed relative reductions of stai ning compared to MAb 3F4 on stored cells, indicating possible structural al terations of PrPC under these conditions.