Fig 1: The pattern of AGR2 staining and specificity(A) The specificity of Rabbit monoclonal anti-human AGR2 antibody to recombinant AGR2 (rAGR2) and AGR3 (rAGR3). 10 ng of each recombinant protein was loaded per lane. Control, 25 mg total protein from a cell lysate from MEF280 cells. Asterisks denote short (*, 1 min) and long (**, 5 mins) exposures. (B) Ishikawa cells were sub-fractionated as described. Whole cell lysate (WCL), cytoplasmic proteins (C), integral membrane and organellular proteins (M) and nuclear and cytoskeletal proteins (N/CS). AIF, apoptosis-inducing factor, is associated with the mitochondria. Subcellular fractionation is representative of three experiments. (C) AGR2 immunofluorescence of Ishikawa cells, in both monolayer and 3D culture. The bright field image is a representative image of Ishikawa cells forming a spheroid 3D culture. Scale bar 50 µm.