FIG 4.

Cell-by-cell analysis of dimerization by confocal microscopic FRET. (A, B) FRET efficiencies of donor (ECFP)- and acceptor (EYFP)-tagged Fos²¹⁵ or full-length Fos molecules as a function of the acceptor-to-donor molecular ratio (N_A/N_D). Data from 300 cells were grouped into three subsets as a function of donor intensity (low, <800; medium, 800 to 1,200; high, >1,200 [arbitrary units]). Cellular data were binned in 0.25-wide intervals of the N_A/N_D values to reduce data scatter. FRET efficiencies increased with increasing N_A/N_D ratios. (C) Saturation values of FRET efficiencies at high N_A/N_D ratios (>0.95). ECFP-EYFP fusion protein served as a positive control, and independently expressed ECFP and EYFP served as a negative control. The FRET data of the Fos-Jun and Fos²¹⁵-Jun pairs were previously published in reference 34.