Approaches to purification and generation of monoclonal antibodies of an insulin-sensitive 90 kd protein
AdvisorSammons, David W.
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PublisherThe University of Arizona.
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AbstractInsulin action has been extensively studied although the exact mechanism by which the binding of this hormone to its receptor causes the observed effects is still obsure. A 90 kd membrane protein may be involved in this mechanism. An attempt was made to purify and make a monoclonal antibody to the 90 kd protein. Several purification methods were attempted and a 2-D electrophoretic procedure developed. Conventional hybridoma production methods were tried as well as a novel hybridoma procedure using selection of J11Dlo cells, culture in LPS/DxSO4, and electrofusion. The resulting monoclonal antibodies to the 90 kd protein were cross-reactive. Furthermore, the immunological results and the presence of the 90 kd protein in several mammalian species suggest that this protein may be evolutionarily conserved and may play a role in insulin action.