Abstract
Platelet activation is traditionally quantified using turbidimetric aggregometry, which reflects integrin αIIbβ3 activity, an important determinant of platelet function during pathophysiological thrombus formation. However, aggregometry does not recreate the shear conditions prevailing during thrombosis in vivo. Here we describe novel whole-frame analysis of real-time video microscopy to quantify platelet adhesion receptor activity under shear in parallel-plate flow chambers. We demonstrate that the rate of change of surface coverage (designated Platelet Population Mobility, PM) quantifies platelet mobility. On collagen, PM falls exponentially to a low level, corresponding to firm platelet adhesion, while on other substrates, PM remains high. Different receptor-specific thrombogenic surfaces reveal that the PM time constant reflects real-time changes in integrins αIIbβ3 and α2β1 activity. This ensemble kinetic analysis has the potential to provide valuable diagnostic information about platelet thrombus formation with both academic and clinical applications.
Acknowledgements
The authors thank Joanna-Marie Howes, Stephanie Jung, Ben Maddox, Masaaki Moroi and Andrew Thompson (University of Cambridge, UK) for useful feedback and discussions of this manuscript.
Declaration of interest
The authors declare no competing financial interests. This work was supported by the British Heart Foundation (RG/09/003/27122), the Medical Research Council (G0500707) and The Newton Trust.
Supplementary material available online
Figure S1. Illustration of the subtraction of subsequent frames to yield a dynamic measure of platelet deposition on a thrombogenic surface.
Figure S2. PM quantifies the activation state of a2b1 and aIIbb3 under flow conditions.
Supplemental material can be viewed and downloaded at http://informahealthcare.com/plt