Production and histological application of affinity-purified antibodies to heat-denatured green fluorescent protein.

ABSTRACT: Enhanced green fluorescent protein (GFP) irreversibly loses not only fluorescence but also antigenicity recognized with conventional anti-GFP antibodies by heat denaturation. This hinders combinatory applications of the GFP immunodetection technique with heat-requiring procedures, such as in situ hybridization histochemistry, antigen retrieval, and Western blot. Here we produced new rabbit and guinea pig antibodies against heat-denatured GFP. The polyclonal antibodies affinity-purified with the antigen column detected a single band corresponding to the molecular size of GFP in Western blot analysis, with mouse brain expressing GFP from the GAD67 locus. By immunofluorescence labeling, the new antibodies detected GFP molecules in heat (> or = 70 degrees C)-treated sections but not in untreat

SUBMITTER: Nakamura KC 

PROVIDER: S-EPMC2430162 | biostudies-literature | 2008 Jul

REPOSITORIES: biostudies-literature