Immunochemical Characterization of Type IV Procollagen from Anterior Lens Capsule

Basement membrane collagen from bovine anterior lens capsules (ALC) was isolated by a non-degradative procedure and characterized. The material was obtained by extracting the ALC with 0.5 {scm} acetic acid, passing the extract through a DEAE-cellulose column, collecting and concentrating the unbound...

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Published inCollagen and related research Vol. 5; no. 3; pp. 233 - 244
Main Authors Brinker, Jane M., Pegg, Marie T., Howard, Pamela S., Kefalides, Nicholas A.
Format Journal Article
LanguageEnglish
Published Germany 01.06.1985
Subjects
Animals
basement membrane
Basement Membrane - immunology
Cattle
Cross Reactions
Enzyme-Linked Immunosorbent Assay
Epitopes - isolation & purification
Immunochemistry
Lens Capsule, Crystalline - immunology
Lens, Crystalline - immunology
mmunochemistry
Procollagen - immunology
Protein Conformation
type IV procollagen
basement membrane
mmunochemistry
enzyme-linked immunosorbent assay
type IV procollagen
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Summary:Basement membrane collagen from bovine anterior lens capsules (ALC) was isolated by a non-degradative procedure and characterized. The material was obtained by extracting the ALC with 0.5 {scm} acetic acid, passing the extract through a DEAE-cellulose column, collecting and concentrating the unbound fraction. Amino acid and carbohydrate analysis, polyacrylamide gel electrophoresis and rotary shadowing electron microscopy showed the purified extract to have the characteristics of basement membrane procollagen. Examination by rotary shadowing revealed the material to consist of type IV procollagen-like tetramers in various degrees of aggregation. When this purified procollagen was injected into rabbits, antibody of high titer and specificity was obtained. The competitive ELISA was then used to demonstrate lack of reactivity of the antibody with collagen types I, II, III and V and fibronectin. The electroimmunoblot technique was used to demonstrate lack of reactivity with laminin. Competition with collagenase resistant fragments of type IV procollagen representing the carboxyl terminal domain (NCI) and the 7-S domain, as well as with a pepsin-resistant α1(IV)125K chain was markedly weaker as compared to the native type IV procollagen molecule.
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ISSN:0174-173X
DOI:10.1016/S0174-173X(85)80013-3