Fig. 5.

Keratanase and sialidase pretreatments diminish Siglec-8 binding to purified Siglec-8 ligands. Siglec-8 ligands were extracted from human trachea, purified by sequential size-exclusion and Siglec-8 affinity chromatography and subjected to keratanase or sialidase treatments. Samples were resolved on 1.5% acrylamide, 2% agarose composite gels, blotted to PVDF membranes and probed with precomplexed Siglec-8-Fc or anti-aggrecan antibody (7D4) as indicated. Lanes: (1) incubation without enzyme; (2) sialidase (67 mU/mL, 90 min); (3) keratanase II (6 mU/mL, 16 h). HiMark molecular weight standards are shown at the left (Stds).