FIG. 3.

Western analysis of various HBMEC fractions with anti-Ecgp antibody. HBMEC membrane proteins were subjected to WGA and MAL affinity chromatography as described in Materials and Methods. The MAL-bound fraction (0.5 mg) was incubated with E44 (E44+MAL-bound) or E91 (E91+MAL-bound), the bound proteins were released and separated by SDS-PAGE. After the proteins were transferred to nitrocellulose, the blot was subjected to immunoblotting with anti-Ecgp antibody. Similarly, deglycosylated Ecgp (Deglyc-Ecgp), MAL-bound either HUVEC (E44+HUVEC), or CEC (E44+CEC) membrane proteins were also analyzed as described. WGA-bound and MAL-bound fractions were also loaded onto the gel to examine the protein pattern. The molecular weight markers are indicated on the left.