Figure 2.

General working principles of force measurements (A) and examples of force distance recordings (B). (A) Chimeric VE-cadherin-Fc is covalently attached by PEG linkers to plate and cantilever tip of the AFM. Cadherins bound to tip and plate by flexible PEG linkers are capable of free equilibrium diffusion and interaction with opposing cadherins during tip-to-plate encounter. Molecules are brought into binding contact by upward movement of the plate. During downmovement the force required for unbinding (f_u) is proportional to deflection to the cantilever (Δz) and its known spring constant. The total length of the stretched PEG-adhesion dimer is defined as unbinding length (l_u). (B) Force-distance curves obtained at a retrace velocity of 800 nm·s⁻¹ and 0.1 s encounter duration in buffer A containing 2 mM Ca²⁺. Examples of single unbinding events and multiple unbinding events are shown. Asterisks mark unbinding events taken for statistical evaluation. Unbinding events were completely abolished by addition of EGTA (5 mM) and antibody to VE-cadherin external domain.