EXPERIMENTAL-STUDY OF THE INTERACTION RANGE AND ASSOCIATION RATE OF SURFACE-ATTACHED CADHERIN-11

We describe a method allowing quantitative determination of the intera ction range and association rate of individual surface-attached molecu les. Spherical beads (1.4 pm radius) were coated with recombinant oute r domains of the newly described classical type hi cadherin 11, a cell adhesion molecule. Beads were driven along cadherin coated surfaces w ith a hydrodynamic force of approximate to 1 pN, i,e,, much less than the mechanical strength of many ligand-receptor bonds. Spheres display ed periods of slow motion interspersed with arrests of various duratio n, particle position was monitored with 50 Hz frequency and 0.025 mu m accuracy, Nearly 1 million positions mere recorded and processed. Com parison between experimental and computer-simulated trajectories sugge sted that velocity fluctuations might be related quantitatively to Bro wnian motion perpendicular to the surface, The expected amplitude of t his motion was of order of 100 nm, Theoretical analysis of the relatio nship between sphere acceleration and velocity allowed simultaneous de termination of the wall shear rate and van der Waals attraction betwee n 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 funct ion 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 ap proximate to 10 nm, It is concluded that the presented methodology all ows sensitive measurement of sphere-to-surface interactions (with appr oximate to 10 fN sensitivity) as well as the effective range and rate of bond formation between individual adhesion molecules.