Luminometric assay of platelet activation in 96-well microplate

Abstract

As of today, no practical method for large-scale functional anti-thrombosis agent screening exists. Based on the phenomenon that platelet activation results in the release of ATP from dense granules, we report the development and optimization of a 96-well microplate luciferase assay to assess platelet activation via luminescence detection of the released ATP. In addition, the assessment of re-calcification-induced clotting of citrated platelet-rich plasma (PRP) is also possible. Collagen, thrombin, U46619, and ADP were shown to induce platelet activation in a concentration- and time-dependent manner. The assay is applicable to PRP, washed platelets, and whole blood. Fundamentally, this is an ideal protocol for screening large numbers of anti-thrombotic drugs because of its sensitivity and the low amount of platelets required to detect simultaneous platelet activation.