Chemolabeling of frozen cerebral tissue proteins and immunopurified products with biotin and digoxigenin: physicochemical characteristics of biotinylated and digoxigeninated products.
Biotinylation and digoxigenination have been compared for labeling proteins from a total frozen tissue extract and from products of immunopurification with anti-RAR beta (retinoic acid receptor beta). The detection of biotinylated and digoxigeninated proteins was found to be easier and more sensitive than detection of silver-stained proteins after two-dimensional electrophoresis. Although biotinylated or digoxigeninated proteins can be detected with avidin conjugates or anti-digoxigenin antibodies, they can also be detected with specific antibodies such as anti-RAR beta antibodies. Previously, coimmunoprecipitates could be visualized only by radioactive amino acid incorporation in cell culture, whereas biotinylation and digoxigenination enable the study of specific protein expression in frozen tissues by immunoprecipitation and the visualization of coimmunoprecipitates. Chemolabeling presents the two major advantages of limiting the use of radioisotopes and allowing the use of frozen tissues in all types of protein expression studies.
Laboratoire de Neurobiophysique, INSERM U318, Grenoble, France.
J F
JF
Brunet
F
F
Berger
G
G
Amalfitano
A L
AL
Benabid
D000418:Albumins / Q000032:analysis; D001710:Biotin / Q000737:chemistry*; D001923:Brain Chemistry; D004076:Digoxigenin / Q000737:chemistry*; D015180:Electrophoresis, Gel, Two-Dimensional; D005615:Freezing; D006801:Humans; D008970:Molecular Weight; D009419:Nerve Tissue Proteins / Q000032:analysis*; D011233:Precipitin Tests
D000418:Albumins; D009419:Nerve Tissue Proteins; D001710:Biotin; D004076:Digoxigenin