Preferential binding of CHO-131 to sialomucins.

(A) Neutrophil staining by CHO-131 is diminished following O-glycoprotease treatment. Peripheral blood neutrophils were subjected to O-glycoprotease (solid line; 48 μg/mL, 5 × 10⁶neutrophils/mL) or sham treatment (filled histograms), and then labeled with the mAbs CHO-131, HECA-452 (anti-sLe^X), or 215 (anti–PSGL-1) at 15 μg/mL. Nonspecific antibody labeling was determined with the use of the appropriate isotype negative control mAbs (dashed lines). Cell staining levels were examined by flow cytometry, and 10 000 cells were examined per sample. Representative data from multiple repetitions are shown. (B) CHO-131 binds immunoprecipitated PSGL-1. Human neutrophil PSGL-1 was immunoprecipitated with the anti–PSGL-1 mAb PL1 and adsorbed to 4-μm latex microspheres, as described in “Materials and methods.” The microspheres were labeled with CHO-131, the anti–PSGL-1 mAbs 215 and PL1, the anti–L-selectin mAb DREG-200, or an anti-CD11a mAb at 5 μg/mL, as indicated (filled histograms). Nonspecific antibody labeling was determined with the use of the appropriate isotype negative control mAbs (solid line). Microsphere staining was assessed by flow cytometry, and 5000 microspheres were examined per sample. Representative data from multiple repetitions are shown.