Fig. 4. Binding of CRP to Fn.

Microtiter wells were coated with Fn. A, increasing concentrations of purified nCRP in TBS, pH 7.0, with and without 5 mM CaCl₂ were added to the wells. B, nCRP, 10 μg/ml in TBS, pH 7.0, containing increasing concentrations of CaCl₂ were added to the wells. Binding of CRP to Fn in the absence of CaCl₂ was taken as 100% to calculate percent inhibition. C, increasing concentrations of purified WT and mutant CRP in TBS, pH 7.0, without CaCl₂ were added to the wells. D, increasing concentrations of purified WT and mutant CRP in TBS, pH 5.0, without CaCl₂ were added to the wells. E, increasing concentrations of purified WT and mutant CRP in TBS, pH 5.0, with and without CaCl₂ were added to the wells. F, increasing concentrations of purified nCRP, 10 μg/ml in TBS at increasing pH, with and without Ca²⁺ were added to the wells. The binding of CRP to Fn in the absence of Ca²⁺ at various pH was plotted as 100%. In all, bound CRP was detected by using polyclonal anti-CRP antibody as the reporter. All experiments were performed at least three times, and a representative experiment is shown.