Experimental study of the interaction range and association rate of surface-attached cadherin 11

We describe a method allowing quantitative determination of the interaction range and association rate of individual surface-attached molecules. Spherical beads (1.4 pm radius) were coated with recombinant outer domains of the newly described classical type hi cadherin 11, a cell adhesion molecule. Beads were driven along cadherin coated surfaces with a hydrodynamic force of approximate to 1 pN, i,e,, much less than the mechanical strength of many ligand-receptor bonds. Spheres displayed periods of slow motion interspersed with arrests of various duration, particle position was monitored with 50 Hz frequency and 0.025 mu m accuracy, Nearly 1 million positions mere recorded and processed. Comparison between experimental and computer-simulated trajectories suggested that velocity fluctuations might be related quantitatively to Brownian motion perpendicular to the surface, The expected amplitude of this motion was of order of 100 nm, Theoretical analysis of the relationship between sphere acceleration and velocity allowed simultaneous determination of the wall shear rate and van der Waals attraction between spheres and surface, The Hamaker constant was estimated at 2.9 x 10(-23) J. The frequency of bond formation was then determined as a function of sphere velocity. Experimental data were consistent with the vie cv that the rate of association between a pair of adhesion molecules was approximate to 1.2 x 10(-3) s(-1) and the interaction range was approximate to 10 nm, It is concluded that the presented methodology allows sensitive measurement of sphere-to-surface interactions (with approximate to 10 fN sensitivity) as well as the effective range and rate of bond formation between individual adhesion molecules.