Simultaneous Detection of Klotho and NEAT1

Slides of HK-2 cells were fixed with 4% paraformaldehyde for 20 min and processed by a combined immunofluorescence and FISH protocol, which was developed for the simultaneous detection of Klotho and NEAT1^{22 (link)}. First, the slides were hybridized with 8 ng/μl NEAT1 probes (Ribobio, Guangzhou, China) at 37 °C overnight. Subsequently, the slides were incubated with anti-Klotho (1:25, Santa Cruz, USA) at 4 °C overnight. Then, the reaction was developed with FITC goat anti-mouse IgG (1:100, ProteinTech, China) for 1 h. Finally, nuclei were counterstained with DAPI. The slides were visualized for immunofluorescence and FISH with a fluorescence microscope at a magnification of ×400.