Chemical Characterization of N-Linked Oligosaccharide As the Antigen Epitope Recognized by an Anti-Sperm Auto-Monoclonal Antibody, Ts4
Fig 7
Western blot analysis of the testicular TS fraction after endo- or exo-glycosidase digestion.
Aliquots containing 20 μg of testicular TS extract were digested with various kinds of endo- or exoglycosidase. After the reaction mixture was incubated for 24 h at 37°C, 5 μg of proteins was applied to each lane of the 10% SDS-PAGE gel under reducing conditions and then was probed with the Ts4 mAb (upper panel) or the 6035 mAb (lower panel). Lane 1: N-glycanase (0.1 unit/ml), lane 2: endoglycosidase F1 (0.1 unit/ml), lane 3: endoglycosidase F2 (0.05 unit/ml), lane 4: endoglycosidase F3 (0.05 unit/ml), lane 5: endoglycosidase H (0.05 unit/ml), lane 6: in absent of endoglycosidase, lane 7: α-mannosidase (7 unit/ml), lane 8: β-mannosidase (0.03 unit/ml), lane 9: β-N-acetylglucosaminidase (2 unit/ml), lane 10: neuraminidase (0.16 unit/ml), lane 11: α-L-fucosidase (0.6 unit/ml), lane 12: in absent of exoglycosidase.