This invention discloses a live-
cell drug screening method and
system based on FRET two-
hybrid analysis. Firstly, preliminary experiments are conducted on multi-well plates using
cell samples transfected with FRET standard plasmids to determine experimental conditions, including
cell seeding density, imaging objective, excitation
light intensity,
field of view analysis range, and measurement of FRET
system calibration parameters. Secondly, live-cell
drug screening based on FRET two-
hybrid analysis is performed under the preliminary experimental conditions.
Cell culture is first performed, and
drug and control groups are set up, followed by FRET three-channel imaging to calculate E0. D E A and R c And draw E D -R c Figure and E A -(1 / R) c (Figure); then use E D -R c With E A -(1 / R) c The linearly separated FRET two-
hybrid analysis method yielded E Dmax E
Amax and N A / N D Data was screened using the
saturation index sRatio; finally, the active and potential active substances in the
drug group were obtained through a
significant difference test. This application enables accurate and rapid quantitative FRET measurement under a wide-field
microscope with low
magnification, meeting high-
throughput requirements and screening for active compounds in live cells.