Figure 2

Subcellular relative distribution of APL^*-target species and effect of EGCG 100 μM pre-treatment from Phasor analysis of the FLIM images. Phasor color maps of FLIM XY sections, determined at CH1 (Ph ₁ ) and CH2 (Ph ₂ ), no treated (-EGCG; first column), and pre-incubated with EGCG 100 μM for 30 minutes (+EGCG; second column) previous to [APL^*] addition, of representative groups of (a) HeLa-wt, and (b) HeLa-APL-R cells, treated with [APL^*] 5 nM for t ∼ 20 minutes, superimposed on a general grayscale intensity image of the whole group of cells. Phasor Plots (third column) showing the changes in the phasor cluster location associated with the formation of the different APL^*-target species at different times, identified in the phasor plot with the corresponding color cursors. Cellular regions showing the maximum fractional contribution of APL^*-B complexes at CH1 (Ph ₁ ; blue scale cursors); cellular regions in which coexist with similar fractional intensities APL^*-A and APL^*-B complexes at CH2 (Ph ₂ ; green scale cursors); and cellular regions showing the maximum fractional contribution of APL^*-A complexes at CH2 (Ph ₂ ; brown scale cursors). Light to dark colors indicate low to high total APL^* fluorescence intensity pixels, high to low fractional contribution of the AF to the total fluorescence intensity, with similar fractional contributions to the total pixel intensity. Black lines in the phasor plots represent the linear combination of 100% pure APL^*-A complexes and autofluorescence AF (A-AF₁ at CH1, and A-AF₂ at CH2), and 100% pure APL^*-B complexes and autofluorescence AF (B-AF₁ at CH1, and B-AF₂ at CH2). λ_exc = 750 nm. Channel #1: FF01 520/35, Channel #2: FF02 435/40, Dichroic filter: FF458-Di02. 1.2 ms/pixel.