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Fig. 6. The bond strength between recombinant cadherin dimers increases with increasing contact time. (A) AFM cantilevers coated with recombinant OB-cadherin (10 µg/ml) were put into contact with similarly coated surfaces, using a constant approach velocity (0.1 µm/second) and loading force (3 nN). All measured rupture forces are summarized in histograms that are fitted with Gaussian curves for contact times of 2, 5, 10 and 60 seconds (n
5000 in each condition). Gaussian curves obtained after 5, 10, and 60 seconds contact time are normalized to the total number of events obtained after 2 seconds of contact time for direct comparison. The dashed lines represent Gaussian curve fits of single peaks that are included in each total data set. (B) Overlaying all Gaussian fits obtained for each contact time demonstrates increasing formation of a third force peak with increased contact time, at decreasing amplitude of the first peak. Inset in B shows the average number of rupture jumps (±s.d.) that precede complete separation of two recombinant OB-cadherin bonds after different contact times. (C) AFM cantilevers coated with 10 µg/ml recombinant OB-cadherin were put into contact with surfaces that exhibited recombinant OB-cadherin coatings in decreasing concentrations of 20, 10, 8, 5, 2, and 1 µg/ml (n5000 in each condition), using a constant approach velocity (0.1 µm/second), loading force (3 nN) and a contact time of 60 seconds. Inset in C shows the average number of rupture jumps (±s.d.) that precede complete separation of two recombinant OB-cadherin bonds as a function of cadherin density. Note that lowering cadherin concentration does not reduce the number of force peaks but rather decreases the average number of rupture events leading to complete bond separation.
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