The platelet adhesion cascade takes place in the presence of shear flow, a factor not accounted for in conventional (static) well-plate assays. This article reports on a platelet-aggregation assay utilizing a microfluidic well-plate format to emulate physiological shear flow conditions.
While the methods here were presented using a specific extracellular matrix protein and a specific inhibitor of platelet aggregation, the protocol is extensible to other coatings, other cell types and other inhibitors for cell adhesion assays. The key ingredients to success of such experiments are avoidance of bubbles in the microfluidic channels, correct dilution of all reagents in the correct diluents, and appropriate incubation conditions for all reagents.
The authors have nothing to disclose.
Material Name | Type | Company | Catalogue Number | Comment |
---|---|---|---|---|
collagen I, Bovine | Reagent | Invitrogen | A1064401 | other sources of collagen I can be used |
PBS plus Ca/Mg | Reagent | Invitrogen | 14040-117 | |
Bovine serum albumin (BSA) | Reagent | EMD | EM-2930 Bottom of Form | From VWR |
calcein AM | Reagent | Invitrogen | C1430 | Calcein AM from BD can also be used |
BioFlux 1000 System | Reagent | Fluxion Biosciences | Contact Company | |
BioFlux Plate | Reagent | Fluxion Biosciences | 900013 | |
antiGPIIb/IIIa | Reagent | Abcam | ab33407 | an alternative # is ab15021 |
ReoPro | Reagent | Eli Lilly | by Rx only |