Figure 3
From: Implantation Serine Proteinases heterodimerize and are critical in hatching and implantation
Purification of the uterine ISP enzyme complex. A- FPLC chromatogram displaying the affinity purification of ISP1-ISP2 complex from mouse uterine homogenates using a Benzamidine Sepharose column. The inset shows the Western Blot results of this partially purified fraction (lane 1: ISP1-ISP2 complex probed with anti-ISP1 antibody, lane 2: ISP1-ISP2 complex probed with anti-ISP2 antibody). B- FPLC chromatogram displaying the separation of ISP1-ISP2 complex using a Superdex™-200 column. The inset shows the Western blot results of the fraction purified from mouse uterine homogenates by ion exchange chromatography (DEAE-Sepharose) followed by gel filtration using Superdex™-200 and Superdex™-75 columns (lane 1: ISP1-ISP2 complex probed with anti-ISP1 antibody, lane 2: ISP1-ISP2 complex probed with anti-ISP2 antibody). C- Affinity purification of ISPs by immunoprecipitation (1- eluted ISP1 fraction pulled down by anti-ISP1 antibody covalently bound to acrylamide activated agarose beads in a chromatography column, 2 – eluted ISP2 fraction pulled down by anti-ISP2 antibody covalently bound to acrylamide activated agarose beads in a chromatography column). D- Co-immunoprecipitation of ISPs (1- Fraction from section C1 probed with anti-ISP2 antibody, 2- Fraction from section C2 probed with anti-ISP1 antibody.