Optimization of Protocol for the Study of Glucose Transporters

UNCP Author/Contributor (non-UNCP co-authors, if there are any, appear on document)
Bradley Setser (Creator)
The University of North Carolina at Pembroke (UNCP )
Web Site: http://www.uncp.edu/academics/library
Harold Maxwell

Abstract: The purpose of this experiment was to determine if it was possible to make changes in the current method of glucose transporter study, which is often undertaken in an attempt to find a link between glucose transporters and diabetes. In order to do this, an antibody against rat glut-4, one of the isoforms of glucose transporters was obtained from East Carolina University School of Medicine. The antibody was then purified with a Protein A column. The glut-4 glucose transporter was isolated from homogenized rat skeletal muscle. A liver sample was also taken as a negative control. The proteins take from the homogenization of liver and muscle were immobilized on nylon paper by dot blotting. A primary antibody, antiglut-4 antibody from East Carolina University, was added to the immobilized proteins. A secondary antibody, goat anti-rabbit antibody, which was conjugated to horse radish peroxidase enzyme was added to the protein-primary antibody complex. When substrate for the enzyme was added, a color change would have indicated whether or not the glucose transporter was present. In this experiment, no color change was detected. An ELISA test was performed in an attempt to determine which step in the procedure was not working. Both the primary and secondary antibodies were found to be functional by the ELISA test. This indicates that the problem in the procedure may be in the extraction of the antigen (glucose transporter).

Additional Information

Language: English
Date: 1991
Glucose Transporter, Rat Glut-4, Antibody, Goat Anti-Rabbit Antibody, Horse Radish Peroxidase Enzyme, ELISA test, Antigen,

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