DIRECT MEASUREMENT OF PLATELET - COLLAGEN INTERACTION BY AFFINITY CHROMATOGRAPHY ON COLLAGEN-SEPHAROSE

  • 1 January 1976
    • journal article
    • research article
    • Vol. 87  (3) , 525-534
Abstract
A method for studying the platelet:collagen interaction is described that permits simultaneous measurement of platelet adhesion to collagen and the collagen-initiated release reaction. Plasma-free platelets were passed through short columns composed of polymeric collagen covalently linked to agarose (Sepharose 2B). EDTA (0.3 mM) was used to prevent platelet aggregation. 14C-Serotonin was used to measure the extent of the release reaction. Measurement of adhesion was based upon 51Cr. The extents of both adhesion and serotonin release were a function of the total collagen content of the column and of the number of platelets applied. Up to 100% of the applied platelets adhered to the columns. As much as 70% of the platelet serotonin was released. Intracellular 51Cr, lactate dehydrogenase and pyruvate kinase were not lost from the platelets. Plasma-free platelets were prepared by 2 different techniques: gel filtration and differential centrifugation. Both preparations gave the same results. At temperatures below 37.degree. C., there was a sharp drop in serotonin release, but only a slight decline in platelet adhesion to collagen. The collagen/Sepharose assay system should provide a usable and greatly needed technique for studying the molecular basis for the platelet:collagen interaction for assessing platelet function in abnormal states and for investigating the mechanism of action of potential inhibitors.

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