Quantitative western blot analysis and spot immunodetection using time-resolved fluorometry.

E P Diamandis; T K Christopoulos; C C Bean

doi:10.1016/s0022-1759(12)80015-9

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Quantitative western blot analysis and spot immunodetection using time-resolved fluorometry.

E P Diamandis·T K Christopoulos·C C Bean

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Abstract

We describe a new method of staining and quantification of proteins blotted or spotted on nitrocellulose. Blotted or spotted proteins are first reacted with specific antibodies followed by reaction with biotinylated secondary antibodies. The immunocomplex is then reacted with a streptavidin-based macromolecular complex labeled with the fluorescent europium chelate of 4,7-bis(chlorosulfophenyl) 1,10-phenanthroline-2,9-dicarboxylic acid (BCPDA). The fluorescent spots or bands can then be assessed by visual inspection under UV illumination, by instant photography or quantified by scanning with a time-resolved fluorometer. The method does not involve enzyme detection, is simple, sensitive and gives sharp bands which remain fluorescent for long periods of time (months to years).

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Animals Blotting, Western Carcinoembryonic Antigen Chelating Agents Densitometry Electrophoresis, Polyacrylamide Gel Europium Ferritins Fluorescent Dyes Fluoroimmunoassay Humans Phenanthrolines Proteins alpha-Fetoproteins Journal Article

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E P Diamandis, T K Christopoulos, C C Bean. 1992-03-04. Quantitative western blot analysis and spot immunodetection using time-resolved fluorometry.. https://doi.org/10.1016/s0022-1759(12)80015-9

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