Abstract
Rapid advances in non-fouling surface technology have pushed the performance of novel coatings toward the detection limit of established protein density quantification techniques. Hence, there is an urgent need for more sensitive detection strategies. Previously we demonstrated that landing rate measurements of microtubules can reveal kinesin surface coverages between 0.1 and 10 μm−2. In this report, we quantify the binding kinetics of highly fluorescent markers to surface-adhered proteins and demonstrate the of protein surface densities in the range of 0.1–1000 μm−2. We utilize this technique to measure kinesin densities on casein-coated glass surfaces and fibrinogen densities on non-fouling polyethylene glycol methacrylate (PEGMA) surfaces. The use of nanospheres (i) potentially permits the detection of a variety of adsorbed proteins, (ii) facilitates the determination of the landing rate due to their uniformity, and (iii) extends the dynamic range of the method due to their small size.
