Cell unroofing setup and process. (A) Diagram of the experimental setup used to unroof cells in a chamber on the microscope stage (see Materials and methods). (B) A field of intact HEK293T/17 cells transiently transfected with TRPV1-YFP and imaged with bright field, epifluorescence (EPI), or TIRF microscopy, as indicated. (C) The same field of cells as in B after the cells were unroofed. The scale bar shown in the left panel of B is 10 µm and applies to all images in B and C.