Extracellular secretion and subcellular localization of the ECRG4 protein. a) Western Blot analysis with an anti-GFP antibody of concentrated cell culture medium of ECRG4-eGFP transfected HCT116 cells shows that ECRG4 is secreted. 1 = concentrated medium of non-transfected cells; 2 = total lysate of eGFP transfected cells; 3 = concentrated medium of ECRG4-eGFP transfected cells. Arrows point to three different processed forms of the ECRG4-eGFP fusion protein. b) Fluorescence microscopy analysis of HCT116 cells transfected with eGFP, ECRG4-eGFP and GalT-GFP vectors. Note that transfection with the eGFP control vector leads to homogenous staining, whereas cells transfected with the ECRG4 fusion gene show staining of discrete structures. These structures are also stained after transfection of the GFP tagged Golgi marker galactosyltransferase (GalT-GFP).