Figure 1. Simultaneous Permeabilization and Fixation Preferentially Retains the Liganded Glucocorticoid Receptor.
(A) 3617 mouse mammary adenocarcinoma cells expressing a GFP-tagged glucocorticoid receptor (GR) were incubated in the absence or presence of the GR ligand corticosterone for 15 min. Cells were then kept on ice (Live), fixed with 2% paraformaldehyde in fluorescence-activated cell sorting (FACS) buffer (Fix only), or fixed with 0.5% Triton X-100 and 2% paraformaldehyde in FACS buffer (Perm-fix). Cells were washed and GFP-GR quantified by flow cytometry. Histograms are representative of two independent experiments.
(B) Relative increase in GR content of GFP-GR 3617 cells upon addition of corticosterone, after perm-fix with various buffers. Data are geometric means of two independent experiments ± geometric SD.
Cort, corticosterone; MFI, median fluorescence index; Triton, Triton X-100.
See also Figure S1.