Immunoblotting of serglycin core protein from secreted and intracellular HUVEC proteoglycans.
(A) The core proteins from the chondroitinase-digested proteoglycans from HUVEC-conditioned medium and an equal amount of non–cell-conditioned medium were fractionated on DEAE-Sephacel to separate the glycosylated core proteins from chondroitinase enzyme and other contaminating proteins in the carrier albumin. The DEAE-Sephacel fractions were electrophoresed by SDS-PAGE and transferred to nitrocellulose. The blots were immunostained with antiserglycin IgY. (B) Identification of intracellular serglycin core protein by35S-sulfate labeling and immunoblotting. Lanes 1 and 2: Chondroitinase-digested cellular proteoglycans were subjected to SDS-PAGE and Western blotting. The blot was subjected to autoradiography. Lanes 3-4: Undigested proteoglycans were electrophoresed in duplicate. One aliquot was transferred to nitrocellulose for Western blotting, and the other was subjected to autoradiography in the gel.
