Mouse Anti-integrin αIIbβ3 Recombinant Antibody (clone 10E5) (CAT#: PABL-247)

Figure 1 Effects of swapping of murine sequence in β3Hu on 7E3 binding.

(A) Adhesion to immobilized fibrinogen of CHO cells transiently expressing β3Hu or β3Hu–M177-184 in association with human αIIb. Cells were preincubated with 2 mM Ca2+ and different antibodies. Adhesion is expressed as OD550 of solubilized crystal violet-stained cells. Data presented are the mean ± standard deviation of three experiments. (B) Radiograph of immunoprecipitates under nonreduced and reduced conditions of 35S-labeled cells using 10E5 or 7E3. Cells were transiently transfected with β3Hu or β3Hu–M177-184 in association with human αIIb.

Artoni, A., Li, J., Mitchell, B., Ruan, J., Takagi, J., Springer, T. A.,... & Coller, B. S. (2004). Integrin β3 regions controlling binding of murine mAb 7E3: Implications for the mechanism of integrin αIIbβ3 activation. Proceedings of the National Academy of Sciences, 101(36), 13114-13120.

Figure 2 Effects of introducing single murine substitutions in β3Hu C177–C184.

(A) Radioautographs of 35S-labeled cells immunoprecipitated with 10E5 or 7E3 and electrophoresed under nonreduced conditions in SDS/PAGE. (B) Immunoblot analysis using mAb AP3 (anti-β3) of 7E3 or 10E5 immunoprecipitates of lysates of cells transfected with β3Hu or T182N. T182N was immunoprecipitated by 10E5 and migrated as a closely spaced doublet. Only the minor band migrating with the same M r as β3Hu was immunoprecipitated by 7E3. (C) Adhesion to immobilized fibrinogen of 293T cells expressing native and mutated β3. Cells were preincubated with 2 mM Ca2+.

Artoni, A., Li, J., Mitchell, B., Ruan, J., Takagi, J., Springer, T. A.,... & Coller, B. S. (2004). Integrin β3 regions controlling binding of murine mAb 7E3: Implications for the mechanism of integrin αIIbβ3 activation. Proceedings of the National Academy of Sciences, 101(36), 13114-13120.

Figure 3 Effect of substituting alanines for C177 and C184 on 10E5 and 7E3 binding.

Lysates of constructs containing β3Hu or β3Hu–M178-183 and/or C177A+C184A residues were immunoprecipitated by 10E5 (Left) or 7E3 (Right), electrophoresed under nonreduced conditions in SDS/PAGE, and immunoblotted with anti-β3 mAb AP-3.

Artoni, A., Li, J., Mitchell, B., Ruan, J., Takagi, J., Springer, T. A.,... & Coller, B. S. (2004). Integrin β3 regions controlling binding of murine mAb 7E3: Implications for the mechanism of integrin αIIbβ3 activation. Proceedings of the National Academy of Sciences, 101(36), 13114-13120.

Figure 4 Fluorescent intensity of cells expressing αIIbβ3 or αIIb-R320C β3-R563C after incubation with Alexa647-7E3 IgG or Alexa647-10E5, with or without treatment with 2 mM DTT.

Artoni, A., Li, J., Mitchell, B., Ruan, J., Takagi, J., Springer, T. A.,... & Coller, B. S. (2004). Integrin β3 regions controlling binding of murine mAb 7E3: Implications for the mechanism of integrin αIIbβ3 activation. Proceedings of the National Academy of Sciences, 101(36), 13114-13120.