Retention Of Platelets In A Protein A-Sepharose Column As A Measure Of Platelet Associated IgG

Employing the biological property of staphylococcal protein A to bind human IgG a test system has been devised to detect platelet associated IgG (PAIg) by quantitating platelet retention in an agarose column with immobilised protein A.

Platelets were washed free of plasma and resuspended in Tyrode’s buffer with EDTA and 0.1% BSA. Platelet counts were performed on the suspension prior to and following passage through a miniature protein A column and the percentage of platelets retained calculated. An indirect test for serum platelet antibody utilised washed normal platelets following their incubation in patient serum.

Less than 30% normal platelets or normal platelets incubated in normal serum were retained in the affinity column. Similarly in four samples from patients with thrombocytopenia of nonimmune origin there was less than 15% retention in the column. Platelets from two of five patients with probable ITP (all treated with corticosteroids) showed 85% retention in the column. The remaining patients fell within the normal range. The sera from three patients with proven platelet antibody using other techniques produced 65-80% retention in an indirect test. Four other sera from possible ITP patients (with comparatively high platelet counts) showed less than 30% retention. Specificity for PAIg was suggested by inhibition of retention from 80% to less than 30% following pretreatment of the colurrai with purified IgG or normal serum.

The optimum conditions for the technique are being studied and its specificity and sensitivity ascertained in patients with thrombocytopenia of various kinds. If these preliminary results are extended, such a simple technique may prove of diagnostic value.