Comparison of Y-142 and Y-073 biological activities toward sHB-EGF.

A. Binding activity of Y-142 and Y-073 toward sHB-EGF. Anti-HB-EGF antibody was incubated at various concentrations in an sHB-EGF-coated or a BSA-coated plate. Antibody binding was detected with an HRP-labeled anti-mouse IgG antibody. Y-142 binding to BSA, white square; Y-073 binding to BSA, white circle; Y-142 binding to sHB-EGF, black square; Y-073 binding to sHB-EGF, black circle. The data points represent the mean ± standard deviation (SD) of values acquired in duplicate. B. Neutralizing activity of Y-142 and Y-073 against sHB-EGF-induced EGFR phosphorylation. Anti-HB-EGF antibody was incubated at the indicated concentrations in the presence of 10 nM sHB-EGF with NUGC-3 cells for 15 minutes at 37°C. Cell lysates were incubated on an anti-EGFR antibody-coated plate, followed by incubation with HRP-labeled anti-phosphotyrosine antibody. No sHB-EGF, white circle; sHB-EGF and control IgG, white square; sHB-EGF and Y-142, black square; sHB-EGF and Y-073, black circle. The data points represent the mean ± SD of values acquired in triplicate.