Fig. 3. Purification and characterization of a putative receptor for CS chains. (A) Human fibroblast WI38 extracts (lanes 1-4) and chick embryonic fibroblasts (lanes 5 and 6) were subjected to CS affinity chromatography in the presence of Ca²⁺. Cell membrane extracts (lane 1), the flow-through fraction from the CS column (lane 2), the elution fraction with EDTA buffer (lanes 3 and 5) and the subsequent fraction obtained by further elution with 4M urea (lanes 4 and 6) were subjected to SDS-PAGE (10%) and visualized by Coomassie brilliant blue staining. (B) Comparison of the amino-acid sequence of the 68 kDa protein with annexin 6. (C) Proteins in the elution fraction from human WI38 fibroblasts extracts were subjected to SDS-PAGE and western blot analysis. Lane 1, Coomassie brilliant blue staining of SDS-PAGE (10%). Lane 2, identification of immunoreactive human annexin 6 in the elution fraction by western blot analysis with rabbit anti-annexin 6 antibodies. An identical blotting pattern was obtained by analysis using mouse monoclonal antibody to human annexin 6.