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Organelle-specific antibodies: production of antibodies to Golgi subcompartments
J Saraste1, M Bronson, G E Palade
1Department of Cell Biology, Yale University School of Medicine, New Haven, Connecticut 06510.
We have devised a strategy for producing Golgi subcompartment-specific antibodies that involves utilizing Triton X-114-phase separated membrane proteins derived from rat pancreatic Golgi subfractions as immunogens. When we tested the approach by immunizing rabbits with membrane proteins derived from heavy Golgi subfractions that are known to be enriched in cis Golgi elements, we succeeded in generating an antibody that recognized a 58 kD protein that was restricted in its distribution to cis Golgi cisternae in several cell types. Thus we have demonstrated the feasibility of the approach we devised for generation of Golgi subcompartment-specific antibodies, and we have also succeeded in identifying a heretofore unknown cis Golgi marker protein.
We have devised a strategy for producing Golgi subcompartment-specific antibodies that involves utilizing Triton X-114-phase separated membrane proteins derived from rat pancreatic Golgi subfractions as immunogens. When we tested the approach by immunizing rabbits with membrane proteins derived from heavy Golgi subfractions that are known to be enriched in cis Golgi elements, we succeeded in generating an antibody that recognized a 58 kD protein that was restricted in its distribution to cis Golgi cisternae in several cell types. Thus we have demonstrated the feasibility of the approach we devised for generation of Golgi subcompartment-specific antibodies, and we have also succeeded in identifying a heretofore unknown cis Golgi marker protein.