Real time western blot assays utilizing fluorescence resonance energy transfer (fret)

Abstract

A Western Blot assay is performed by performing a probing process on a membrane containing target proteins, by contacting the membrane with a fluorescent resonant energy transfer (FRET) solution and allowing the probing process to proceed for a probing time period. The probing process results in a target protein becoming labeled with both a donor chromophore and an acceptor chromophore, which are effective as a donor-acceptor pair for FRET when so linked to the target protein. While performing the probing process, the labeled target proteins are measured by irradiating the membrane with an excitation light to excite the donor chromophores, wherein in each labeled target protein, the excited donor chromophore transfers energy to the acceptor chromophore by FRET and, in response, the labeled target protein emits an emission light. The intensity of the emission light is then measured. The light measured may be light emitted from the donor chromophore and/or light emitted from the acceptor chromophore.