Characterization of 2 monoclonal IgG2a Abs raised to ERp57. (A) The Western blots of the Abs called Mab1 and Mab2 against human platelet lysate (4 × 10⁸ platelets/lane) and purified ERp57 or PDI (200 μg protein/lane). Differential effect of the 2 IgG2a monoclonal anti-ERp57 Abs on (B) ERp57 activity and (C) lack of cross-reactivity to PDI or (D) ERp5 in the GSSG assay. For these studies, the activity of ERp57, PDI, and ERp5 alone (not shown in curves) was identical to enzyme with 30 μg/mL of the control IgG2a. A total of 30 μg/mL of each Ab was used in these studies except for ERp57 (B) where 10 μg/mL of the inhibitory Mab1 Ab was also tested. A total of 60 μg/mL of Mab 1 did not increase the inhibition (not shown). IgG2a control, ●; Mab1 10μg/mL, ▵; Mab1 30 μg/mL, ▴; Mab2 30 μg/mL, ■). (D) The curves for ERp5 with IgG2a, Mab1, or Mab2 (30 μg/mL of each Ab was used) completely overlapped. Each point is the composite of at least 3 samples using the Di-E-GSSG assay. (B-D) The enzyme was added at ∼ 20 seconds.