Quantitation of specific collagen types from lungs of small mammals

Karen M. Reiser, Jerold A Last

Research output: Contribution to journalArticle

26 Citations (Scopus)

Abstract

A technique for determination of the ratio of Type I to Type III collagen in the lungs of rats and mice is described. Total lung collagen is solubilized by digestion with CNBr. The resultant CNBr peptides are fractionated by chromatography on columns of carboxymethyl-cellulose. One partieular peak from the column contains the Type I collagen-specific peptide α1(I)-CB-8 and its incompletely split precursor, α1(I)-CB-3-8, as well as the Type III-specific peptide α1(III)-CB-8. Fractions containing these peptides are pooled, concentrated, and further purified by electrophoresis on polyacrylamide gels in the presence of sodium dodecyl sulfate. These peptides, of molecular weights 23,000, 36,000, and 11,500, respectively, are well separated by gel electrophoresis. The appropriate gel bands are excised after visualization with Coomassie blue stain: their content of collagenous peptides is quantitated after elution from the gels by determination of hydroxyproline (or proline) content, either colorimetrically or by their contained radioactivity (when labeled collagen is biosynthesized with [3H]proline as a precursor). The described technique is accurate, reproducible, and sensitive enough for determination of collagen types from a single rat or mouse lung. This should be an appropriate method for quantitation of collagen types from any tissue source where a high percentage of the collagen content cannot be solubilized by acid extraction or by treatment with pepsin.

Original languageEnglish (US)
Pages (from-to)87-98
Number of pages12
JournalAnalytical Biochemistry
Volume104
Issue number1
DOIs
StatePublished - May 1 1980

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Mammals
Collagen
Lung
Peptides
Gels
Electrophoresis
Proline
Rats
Carboxymethylcellulose Sodium
Collagen Type III
Pepsin A
Hydroxyproline
Radioactivity
Collagen Type I
Chromatography
Sodium Dodecyl Sulfate
Polyacrylamide Gel Electrophoresis
Digestion
Coloring Agents
Visualization

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

Quantitation of specific collagen types from lungs of small mammals. / Reiser, Karen M.; Last, Jerold A.

In: Analytical Biochemistry, Vol. 104, No. 1, 01.05.1980, p. 87-98.

Research output: Contribution to journalArticle

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