a–c, Rabbit anti-PKD2L1 (Thermo Scientific) recognizes overexpressed PKD2L1. HEK cells were transfected with hPKD2L1-IRES mCherry construct and stained with PKD2L1 antibody. PKD2L1 staining (a) is specific to cells that also express mCherry (b). c, Overlay. d–f, Overexpressed hPKD2L1 localizes to the primary cilium in mIMCD3 cells. mIMCD3 cells were transfected with HA-tagged hPKD2L1 and stained with an anti-HA antibody (d) and anti-acetylated tubulin antibody (e). HA immunoreactivity is visible both in the cytoplasm and in the cilium. f, Overlay. g, h, PKD2L1 antibody labels the primary cilium of mIMCD3 cells. g, Confluent mIMCD3 cells were stained with anti-PKD2L1 antibody used in a; and h, acetylated tubulin antibody to label cilia. PKD2L1 immunoreactivity is visible in the primary cilium. i, Overlay. j–o, Primary MEFs of Arl13b-EGFPtg (j–l) and Arl13b-EGFPtg × Pkd2l1−/− mice (m–o) isolated from E14.5 embryos were stained with anti-PKD2L1 antibody used in a. In Arl13b-EGFPtg MEFs, PKD2L1 immunoreactivity (j) co-localizes with ARL13B–EGFP signal (k) labelling the primary cilium. l, Overlay. m–o, PKD2L1 staining is absent in cilia of Arl13b-EGFPtg × Pkd2L1−/− mice. Scale bars: 10 μm.