Fig. 2. (1-25)-4A1-GFP localises to the trans-Golgi network and this localisation requires the presence of both helix-1 and helix-2. (A) (1-25)-4A1-GFP (green) was transfected into Cos-1 cells. Cells were viewed live (top row) or fixed with 4% paraformaldehyde (middle and bottom rows) and co-stained with either the general Golgi stain, BODIPY ceramide (red), the cis-Golgi marker GM130 or the trans-Golgi marker, TGN46 before visualising, using an Alexa Fluor 594-conjugated secondary antibody (red). (B) Cos1 cells were transfected with constructs encoding (1-25)-4A1-GFP with either helix-1 [2-7(1-25)-4A1-GFP, green, top panel] or helix-2 [14-25(1-25)-4A1-GFP, green, lower panel] deleted. Where indicated, cells were stained with the Golgi marker BODIPY ceramide or the live cell mitochondrial probe, Mitotracker (red). Cells were then visualised live. Merged images are presented as marked and any co-localisation is indicated in yellow. Bars, 10 µm. (C) Quantification of the imaging examples shown in B. Images were analysed using the co-localisation tool of Metamorph Imaging as detailed in the Materials and Methods (n=31 cells; values are mean ± s.e.m.).