FIGURE 4.

Direct binding of HS oligosaccharides with Stab1 and Stab2 ectodomains. Ectodomains/secreted versions of each receptor were immunopurified by V5 resin and were either subject to 5% SDS-PAGE, blotted, and probed with anti-V5 antibody (Lane 1, s190-HARE/Stab2; lane 2, s315-HARE/Stab2; lane 3, sStab1) (A) or incubated with an equal amount of each HS construct for 1.5 h under rotation (B–D). The resin was washed with Tris-buffered saline three times and then mixed with scintillation mixture, and cpm was determined. Counts per minute were normalized to the amount of protein on the resin.