Hc pl apo 20x 0.75 imm corr cs2 objective

The cleared tissues were imaged either by confocal or light-sheet microscopy (Fig. 10, Step 3). The samples were positioned either on a glass slide (Menzel-Glaser, AA00008132E) or in a Petri dish (Thermo Fisher Scientific, S33580A), immersed in immersion oil (RI = 1.51, Thermo Fisher Scientific, NC0586121). Imaging was performed with a Leica SP8 confocal microscope with a HC PL APO 20x/0.75 IMM CORR CS2 objective (Leica Microsystems, 15506343). MHI148-PEI and MHI148-PEI-S were excited at 638 nm and fluorescence was detected using a Cy7 emission filter. For imaging on a light-sheet microscope (Leica Microsystems, Leica TCS SP8 DLS), samples were mounted to a hook-shaped holder, submerged in immersion oil, and evaluated with a HCX PL FLUOTAR 5x/0.15 objective. Images were processed and 3D reconstruction was rendered using Leica LAS X software. All the data acquired were saved as. lif files and read into LAS X. The images from 3D scan data contained in this article are based on the adjustments made in minimum and maximum intensity, shading, gamma and opacity of 3D viewer section in the LAS X software.

Protoplast images were obtained using a Leica TCS SP8 confocal microscope equipped with a HC PL APO 20x/0.75 IMM CORR CS2 objective (Leica Microsystems, Mannheim, Germany), with the exception of images in Supplementary Fig. 7b and Fig. 5a that were taken with a Zeiss LSM880 Airyscan using a 63x/1.4 oil immersion lens. GFP was excited with a White Light Laser (WLL) at 488 nm and the emission detected at 500–550 nm. YFP was excited with a 514 nm laser line and detected at 520–555 nm. mCherry fluorescence was excited at 561 nm and emission was detected between 575–630 nm. Samples, co-expressing two fluorophores were imaged in sequential mode between frames. All images were taken as z-stacks (internal distance is 0.5 um) and further analyses and projections were performed with either ImageJ/(Fiji) software^{88 (link)} or Imaris.
For GUV experiments: Images were acquired by confocal fluorescence microscopy (Nikon Eclipse Ti-E inverted microscope using a Nikon A1R confocal laser scanning system with laser lines: 405 nm, 488 nm, 561 nm, 640 nm; 60x/1.49 oil immersion objective; Nikon Instruments, Inc.) and analyzed using the corresponding NIS Elements software (NIS Elements Confocal 5.20, Nikon Instruments, Inc.) and ImageJ (Fiji win64, Open source).