Panel (A) shows a Western blot of lyasates of: mock transfected Cos-7 cells; RGN transfected cells and rat liver homogenate, as a positive control. The lanes were loaded with 7.5μg of protein and probed with a RGN polyclonal antibody (1:50 dilution). A protein labeled band at approximately 33kDa was observed in RGN-transfected lysate and liver homogenate. The Image was modified for clarity by changing the brightness and contrast as well as some smoothing. Panel (B) shows a fluorescence micrograph of Cos-7 cells transfected with Pcmv6 AC-GFP-RGN, indicating the presence GFP-RGN expressing cells. The white bar indicates 20 µm. (C) Untransfected Cos-7 cells analyzed by flow-cytometry, monitoring both the red (propidium iodide) and green (GFP) channels. The untransfected cells were considered to be fully viable, as all cells were binned within quadrant Q4 (i.e. minimal levels of propidium iodide staining). Panel (D) shows RGN-GFP transfected cells analyses by flow cytometry. The cells were again considered to be fully viable as they were binned within quadrants Q4 and Q3 (minimal levels of PI staining). From the analysis of the green fluorescence channel (GFP) more than 33% of the cells analyzed expressed GFP.