Measuring the time-dependent functional activity of adsorbed fibrinogen by atomic force microscopy

In this work, we measured time-dependent functional changes in adsorbed fibrinogen by measuring antigen--antibody debonding forces with atomic force microscopy (AFM). AFM probes were functionalized with monoclonal antibodies recognizing fibrinogen γ392-411, which includes the platelet binding dodecapeptide region. These probes were used to collect force measurements between the antibody and fibrinogen on mica substrates and the probability of antigen recognition was calculated. Statistical analysis showed that the probability of antibody-antigen recognition peaked at ∼45 min postadsorption and decreased with increasing residence time. Macroscale platelet adhesion measurements on these mica substrates were determined to be greatest at fibrinogen residence times of -45 min, which correlated well with the functional activity of adsorbed fibrinogen as measured by the modified AFM probes. These results demonstrate the utility of this approach for measuring protein function at or near the molecular scale and offers new opportunities for improved insights into the molecular basis for the biological response to biomaterials.