A High Affinity Monoclonal Antibody Recognizing the Light Chain of Human Coagulating Factor VII

Abstract

Factor VII (FVII) is a serine protease-coagulating element responsible for the initiation of an extrinsic pathway of clot formation. Here we generated and characterized a high affinity monoclonal antibody that specifically recognizes human FVII. Recombinant human FVII (rh-FVII) was used for the production of a monoclonal antibody using BALB/c mice. The specificity of the antibody was determined by Western blot using plasma samples from human, mouse, sheep, goat, bovine, rabbit, and rat. Furthermore, the antibody was used to detect transiently expressed rh-FVII in BHK21 cell line using Western blot and sandwich ELISA. A mouse IgG1 (kappa chain) monoclonal antibody clone 1F1-B11 was produced against rh-FVII. The affinity constant (K(aff)) of the antibody was calculated to be 6.4×10(10) M(-1). The antibody could specifically recognize an epitope on the light chain of hFVII, with no reactivity with factor VII from several other animals. In addition, transiently expressed rh-FVII in BHK21 cells was recognized by 1F1-B11. The high affinity as well as the specificity of 1F1-B11 for hFVII will facilitate the affinity purification of hFVII and also production of FVII deficient plasma and minimizes the risk of bovine FVII contamination when fetal bovine serum-supplemented media are used for production and subsequent purification of rh-FVII.

Production of monoclonal antibody against recombinant bovine sex‐determining region Y (SRY) and their preferential binding to Y chromosome‐bearing sperm

“…For the production of mouse anti‐bovine SRY mAbs in ascitic fluid, the Sarial et al method with some modifications was used (Sarial et al., 2012). Briefly, 6 BALB/c mice were primed with 600 μl of 2,6,10,14‐tetramethyl‐pentadecane (Pristane, Sigma‐Aldrich) injected intra‐peritoneal.…”