Resonant Energy Transfer (RET) from an optically excited donor molecule (D) to a non-excited acceptor molecule (A) residing nearby is widely used to detect molecular interactions in living cells. Stoichiometric information, such as the number of proteins forming a complex, has been obtained so far for a handful of proteins, but only after exposing the sample sequentially to at least two different excitation wavelengths. During this lengthy process of measurement, the molecular makeup of a cellular region may change, and this has so far limited the applicability of RET to determination of cellular averages. Here we demonstrate a method for imaging protein complex distribution in living cells with sub-cellular spatial resolution, which relies on a spectrally-resolved two-photon microscope, a simple but competent theory, and a keen selection of fluorescent tags. This technology may eventually lead to tracking dynamics of macromolecular complex formation and dissociation with spatial resolution inside living cells.
@article{arxiv.0810.2341,
title = {Imaging protein interactions in vivo with sub-cellular resolution},
author = {Valerica Raicu and Michael R. Stoneman and Russell Fung and Mike Melnichuk and David B. Jansma and Luca Pisterzi and Michael Fox and James W. Wells and Dilano K. Saldin},
journal= {arXiv preprint arXiv:0810.2341},
year = {2024}
}