Fluorescent Labeling of Several Glycosaminoglycans and Their Interaction with Anti-Chondroitin Sulfate Antibody

The amino groups of aminofluorescein were covalently linked to the carboxyl groups of glycosaminoglycans (GAGs; heparin, heparan sulfate, and the chondroitin sulfates A, B, C, D and E) using 1-ethyl-3-(1,3-dimethylaminopropyl)-carbodiimide and N-hydroxysuccinimide as the catalytic reagents, and fluorescently labeled GAGs were obtained separately. The fluorescently labeled GAGs were transferred onto nitrocellulose-coated glass chips with a manual microarrayer. The retention rate of GAGs on the nitrocellulose membrane after washing was calculated, and their interactions with monoclonal anti-chondroitin sulfate antibody (clone number CS-56) were further studied. The experimental results showed that the retention rates were apparently different between the GAGs and of the order chondroitin sulfate E (51.7%) > chondroitin sulfate A (19.5%) > heparan sulfate (16.4%) > heparin (14.1%) > chondroitin sulfate B (12.3%) > chondroitin sulfate D (10.8%) > chondroitin sulfate C (10.7%). The interaction of GAGs with CS-56 was scientifically compared according to their retention rates on nitrocellulose-carbohydrate chips, and after quantitative calculation, the binding ability of the GAGs to CS-56 is of the order chondroitin sulfate C > D > A > E; furthermore, chondroitin sulfate B, heparin, and heparan sulfate did not show any interaction with CS-56.