Endogenous myoJ is concentrated on contractile vacuole membranes. Shown are wild-type (A–D and I–L) and myoJ-null (E–H and M–P) cells double stained for CV membranes (anti-proton pump in B and F, anti-calmodulin in J and N) and anti-myoJ (purified antibody) (C, G, K, and O). D, H, L, and P show the overlaid images. The black arrows in A, and the corresponding white arrows in B and C, point to phase-lucent CV bladders that are positive for both the proton pump and myoJ. The white arrowheads in B and C point to what may be CV tubules that are positive for both the proton pump and myoJ. We note that although the proton pump stained here is also present in endosomal membranes, its density there is about one tenth that in CV membranes (Gerisch et al., 2002), so at the exposures shown here only CV membranes are visualized. The cells in A–D were overlaid with agar before fixation and imaged in widefield. Bars: 4 µm.