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To test the predominant role of the non-vesicular route for retrograde transport in U-2 OS cells, we inhibited vesicular trafficking using Brefeldin A, which blocks COPI coat formation at Golgi membranes and Wortmannin, which broadly affects endosome formation and trafficking and quantified transport of SM(Y), PE(18:1/Y), PC(Y:16:0), PC(Y/20:4) using the lipid imaging pipeline.