FIGURE 1.

Lubricin binds to recombinant human L-selectin/Fc chimera. A, RA lubricin samples (RA1 and RA2) were separated by SDS-AgPAGE under non-reducing conditions. Recombinant L-selectin/Fc chimera bound to lubricin, whereas sialidase A treatment abolished this interaction (RA1). SF lubricin and L-selectin/Fc chimera were detected by mAb 13 and goat anti-human IgG (Fc-specific) antibodies, respectively. Recombinant lubricin (Re lub) was also analyzed in the same way. No L-selectin was found bound to recombinant lubricin in comparison with native lubricin (RA3). B, immunoprecipitation was verified by silver stain and lubricin-specific antibody (mAb 13). In breakthrough, no L-selectin/Fc chimera was detectable by silver stain. In elutant, eluted L-selectin/Fc chimera could be detected by silver stain, whereas lubricin was detectable by immunoblot (WB). The arrow indicates synovial lubricin, and the arrowhead indicates L-selectin/Fc chimera. C, inhibition ELISA with synovial lubricin and L-selectin. 96-well microtiter plates were coated with L-selectin. Synovial lubricin with or without inhibitors (fetuin, porcine gastric mucins (PGM), or 6-sulfo Le^x (sulfo Lew x)) is shown. Bound lubricin was detected by mAb 13. Treatment with 1% BSA instead of lubricin was used as negative control (Ng).