Quantitation of type I and III collagens using electrophoresis of alpha chains and cyanogen bromide peptides.
The methods of quantitating the relative amounts of type I and III collagens in samples containing crosslinked collagen chains were evaluated using electrophoresis of alpha chains and cyanogen bromide peptides. The densitometry areas of the alpha I(I) chains from type I collagen and the alpha I(III) chains from type III collagen were reduced because of the failure of the crosslinked chains to dissociate. However, the ratios of the unit densitometry areas of these chains (area of chain/micrograms type I or III collagen loaded) were constant for type I and III collagens prepared from the same samples of tissue. A calibration factor, which was the same for dermis and mitral valve, was derived to convert the densitometry area ratios to the weight ratios of type I to III collagens. In contrast, the densitometry areas of the alpha I(I) CB8 (type I collagen marker) and the alpha I(III) CB5 (type III collagen marker) were not reduced by crosslinked collagen chains. A calibration factor was also derived to convert the ratios of the densitometry areas of the marker peptides to weight ratios of type I to type III collagens. Almost identical results were obtained when electrophoresis of alpha chains and of cyanogen bromide peptides was used with these calibration factors to quantitate the relative amounts of type I and III collagens in tissue extracts which contained different amounts of crosslinked chains.
D002675:Child, Preschool; D003094:Collagen / Q000032:analysis* / Q000145:classification; D003488:Cyanogen Bromide; D003720:Densitometry; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008297:Male; D008875:Middle Aged; D010446:Peptide Fragments / Q000032:analysis; D012867:Skin / Q000032:analysis
D010446:Peptide Fragments; D003094:Collagen; D003488:Cyanogen Bromide