Jan 15, 1990

Isolation of bovine type X collagen and immunolocalization in growth-plate cartilage

The Biochemical Journal
T Kirsch, K von der Mark

Abstract

Type X collagen was extracted with 1 M-NaCl and 10 mM-dithiothreitol at neutral pH from fetal-bovine growth cartilage and purified to homogeneity by using f.p.l.c. gel filtration on a Superose 12 column, followed by ion-exchange chromatography on a Mono Q column. The purified protein migrates in SDS/polyacrylamide gels with an apparent Mr of 58,000 under reducing conditions and as a high-Mr oligomer in its unreduced form. The amino acid composition is similar to the published composition of chick type X collagen. Pepsin digestion at 4 degrees C decreases the Mr of the monomer to 43,000; purified bacterial collagenase digests most of the molecule, leaving a non-collagenous domain of apparent Mr 15,000, which probably represents the C-terminal globular domain. The IgG fraction from a rabbit antiserum raised against purified bovine type X collagen was specific for this collagen by the criteria of e.l.i.s.a. and immunoblotting after immunoabsorption with collagen types I, II, IX and XI. Immunofluorescence localization of type X collagen in sections of fetal-bovine and human cartilage was possible after acetone fixation of sections and hyaluronidase treatment. Type X collagen was restricted to the zone of hypertrophic and calcified ...Continue Reading

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Mentioned in this Paper

Immunofluorescence Assay
Immunoblotting, Reverse
Hypertrophic Cardiomyopathy
Immune Sera
Chromatography, DEAE-Cellulose
Zyderm
Polyglobin
Bos indicus
Molecular Sieve Chromatography
Enzyme-Linked Immunosorbent Assay

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