U mniba2 gfp filter cube

Ca²⁺ and pH imaging were performed as previously described [33 , 34 (link)], with some modifications. In brief, isolated taste cells were plated on poly-D-lysine coated coverslips at room temperature. After 30 minutes, cells were loaded with the intracellular pH indicator pHrodo Red AM and calcium indicator Fura-2 AM, using PowerLoad concentrate according to the manufacturer’s instructions (Thermo Fisher). YFP-expressing taste cells were identified using a U-MNIBA2 GFP filter cube (Olympus).. pHrodo Red fluorescence intensity for each cell was measured in response to pH 5.0 solutions buffered with Homo-PIPES (145 mM NaCl, 5 mM KCl, 10 mM Homo-PIPES, 2 mM CaCl₂, 1 mM MgCl₂, 20 mM D-glucose) or with acetic acid (145 mM NaCl, 5 mM KCl, 10 mM acetic acid, 2 mM CaCl₂, 1 mM MgCl₂, 20 mM D-glucose). pHrodo Red was excited with 560 nm light and emission at 630 nm was detected using a U-N31004 Texas Red/Cy3.5 filter cube (Chroma Technologies), while Fura-2 AM was excited by with light of 340 nm and 380 nm and emission at 510 nm was detected with U-N71000aV2 FURA2 WM filter (Olympus). Images were acquired on a Hamamatsu digital CCD camera attached to an Olympus IX71 microscope using Simple PCI software. The pHrodo Red fluorescence intensity of each cell was normalized to its baseline fluorescence in pH 7.4 solution (F₀) before the first acid application to give F/F₀.