Colocalization of exogenous VWF with platelets adhered on collagen-immobilized surface. FITC-labeled VWF or nonlabeled VWF was added to platelet-reduced blood labeled with mepacrine (5 × 104 platelets/μL) to give a final concentration of 2.6 VWF:RCo U/mL and perfused at 1600 s–1. Blood flow was stopped at 2 minutes after start of the flow, and the cover slips were washed and fixed with paraformaldehyde solution. The fluorescent images of FITC and mepacrine were separately obtained by using B and BV filters, respectively. (A) An illustration of the collagen-coated glass plate in the flow chamber. Gray circle and bold arrows indicate the collagen-coated area and flow direction, respectively. (B) Fluorescent images of FITC-labeled VWF on the collagen surface at 3 sites: upper, middle, and lower areas of the flow, as shown in panel A. Mepacrine nonlabeled platelet reduced-blood was perfused after addition of FITC-labeled VWF. (C) Fluorescent images of mepacrine-labeled platelets on the collagen surface at 3 sites: upper, middle, and lower areas of the flow, as shown in panel A. (D) Fluorescent images of the same microscope field of the same sample using mepacrine and FITC fluorescence. Magnification, × 100 for panels B and C, and × 400 for panel D.
