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. Author manuscript; available in PMC: 2014 May 10.
Published in final edited form as: Chembiochem. 2013 Apr 12;14(7):10.1002/cbic.201300029. doi: 10.1002/cbic.201300029

Figure 1. Schematic representation of Me-GFP-PABP1 TR-FRET assay for detecting CARM1 cellular activity.

Figure 1

MCF7 cells are infected with BacMam virus encoding a GFP-PABP1 fusion protein and plated in a 384-well assay plate. 24 hours after infection, cells are lysed and incubated for 1 hour in the presence of Me-PABP1 specific primary antibody (Me-PABP1 Ab) and goat anti-rabbit IgG chemically labeled with a terbium chelate (Tb-2nd Ab). After incubation, Tb is excited at 340 nm, and TR-FRET emission ratio of 520 nm over 495 nm is obtained. The association between the antibody and Me-GFP-PABP1 allows energy transfer to occur between the excited-state donor fluorophore Tb and acceptor fluorophore GFP, leading to an increase of the TR-FRET ratio. TR-FRET ratio obtained with Tb-2nd Ab alone is subtracted as background signal.