Immunological characterization of the major chick cartilage proteoglycan and its intracellular localization in cultured chondroblasts: a comparison with Type II procollagen.

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Abstract
Polyclonal antibodies were raised in a rabbit against the major proteoglycan of chick sternal cartilage. A total of 6 antisera was obtained, 3 after the 1st booster injection (A1, A2 and A3) and 3 after the 2nd booster injection (A4, A5 and A6). The A1 antiserum, which was characterized in most detail, immunoprecipitated native as well as chondroitinase ABC-digested or chondroitinase, ABC/keratanase-digested cartilage proteoglycan synthesized by cultured chick chondroblasts, but failed to immunoprecipitate the major proteoglycan synthesized by chick skin fibroblasts. This antiserum was also able to immunoprecipitate the cartilage proteoglycan core protein newly synthesized by cultured chondroblasts, but no other major cell protein. However, the late bleed antisera obtained from the same rabbit after a 2nd booster injection reacted with a new chondroblast-specific polypeptide(s) of .apprx. 60,000 MW in addition to the cartilage proteoglycan. By immunofluorescence procedures, the A1 antiserum stained the extracellular proteoglycan matrix of cultured chondroblasts but not that of skin fibroblasts. Following enzymatic removal of the extracellular matrix and cell membrane permeabilization, this antiserum stained primarily a large, juxtanuclear structure. Additional radioautographic evidence suggests that this structure represents the Golgi complex. Similar immunofluorescence staining with antibodies to the cartilage-characteristic type II collagen revealed that type II procollagen was localized in numerous cytoplasmic, vacuole-like structures which were scattered throughout most of the chondroblast cytoplasm but were notably scanty in the Golgi complex area. The transit of the major cartilage proteoglycan through the Golgi complex of cultured chondroblasts and possible differences in the intracellular distribution of naewly synthesized cartilage proteoglycan and type II procollagen are suggested.