Date of Award

1978

Degree Type

Thesis

Degree Name

Master of Science in Pharmacology and Toxicology

Department

Pharmacology and Toxicology

First Advisor

George C. Fuller

Abstract

Purified collagen were made fluorescent by coupling with 2-methoxy-2,4-diphenyl-3(2H)-furanone, MDPF, and the fluorescent products were separated by sodium dodecyl sulfate- polyacrylamide gel electrophoresis, SDS-PAGE. The profiles of Coomassie blue stained and MDPF-labeled collage. Tl bands on the gels were si."nilar except .for a slight increase in mobility with the MDPF coupled protein. · The relationship between the amount and the area under the peaks recorded from fluorometric scanning of purified al(I) ,a2 and al(III) was linear from 10-5g to l0- 8g. The standard curves for all three a chains were similar. Results from non-replicate determinations had an experimental error of + 6% SE. A mixed sample of cyanogen bromide (CB) peptides from type I and III collagen were quantitated by measuring the peak areas of known peptides from each type. The quantitation of collagen by the coupling of MDPF before electrophoresis is an improvement over staining with Coomassie blue after electrophoresis; since it provides a wider range of linearity, greater sensitivity, peak area independent of distance migration and less variability. The fluorescent method (MDPF-SDS-PAGE) also permits observation of bands during electrophoresis and quantitation immediately after electrophoresis.

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