Extended Data Fig. 3: Calculated subtraction of bleedthrough from EPYC1-GFP to the MITH1-mCerulean channel.

a, EPYC1-GFP bleeds into the mCerulean channel, as seen by the mCerulean signal present in the CrRBCS2;EPYC1 line despite the absence of MITH1-mCerulean. b, Original images of the CrRBCS2;EPYC1-GFP;MITH1-mCerulean line. c, CrRBCS2;EPYC1-GFP;MITH1-mCerulean images following subtraction of a constant value for background signal in the GFP and mCerulean channels. d, CrRBCS2;EPYC1-GFP;MITH1-mCerulean images following subtraction of calculated bleedthrough of GFP signal into the mCerulean channel. e, CrRBCS2;EPYC1-GFP;MITH1-mCerulean images after further denoising and deconvolution as detailed in Methods. f, Scatter plot displaying the association between GFP signal intensity and mCerulean intensity in the CrRBCS2;EPYC1-GFP line (not expressing mCerulean); each dot represents the average mCerulean and GFP signals from a region in an EPYC1-GFP condensate. The slope of the regression was used to quantify the amount of bleedthrough from GFP into the mCerulean channel. g, Additional information on the calculated bleedthrough signal. From left to right, the mCerulean channel image from panel c is shown, followed by the calculated bleedthrough signal from the GFP channel into the mCerulean channel, followed by the image resulting from the subtraction of the calculated bleedthrough signal (this image corresponds to the image shown in the mCerulean channel of panel d).