Fluoview fv3000 microscope

Immunostaining was conducted to find the expression of caspase-3 protein in normal hepatocytes, CCl₄-treated and compounds-treated, MSCs-treated and MSCs + compounds-treated hepatocyte. First, the cells were washed 3 times with PBS, each for 5 min, fixed in 4% formaldehyde for 30 min at room temperature and permeabilized with 0.1% triton for 10 min; then, nonspecific binding was inhibited by blocking with 2% BSA in PBS for 45 min. The cells were then incubated with primary antibodies for 4 h at room temperature. The primary antibody used for caspase-3 was rabbit monoclonal (1:1000, abcam) and the secondary antibody was goat anti-rabbit (1:500, abcam, Cambridge, UK). The samples were incubated for 45 min at room temperature (RT), and then were washed with PBS and incubated with DAPI for 2–5 min at room temperature. Then, the samples were again washed with PBS, and subsequently mounted with vecta sheets. The samples were then observed using a Fluoview FV 3000 microscope (Olympus, Tokyo, Japan) and pictures were taken.

We placed mated females from representative Uhg4 deletion lines and their controls (208-ΔF, 208-ΔG, DGRP_208) in fresh food vials supplemented with yeast paste every 12 h for 36 h prior to dissection. Flies were dissected in 1X PBS and ovarioles were gently separated. Ovaries were fixed in 4% paraformaldehyde for 15 min, followed by three 15-min washes in PBS with 0.2% Triton X-100. Following a final 15-min wash in PBS, ovaries were stained with DAPI (Invitrogen) (1 μg/mL) for 10 min and mounted with ProLong Gold (Invitrogen) immediately after the final PBS wash. Ovaries were imaged on an Olympus Fluoview FV3000 microscope at 20 × magnification. Images were processed in Fiji [102 (link)].

The absorbance spectra of UCNPs, DPP, ICG, UCNPs-D, UCNPs-I, and UCNPs-DI under various conditions were measured by an ultraviolet/visible absorption spectrometer (Lambda-35 UV/visible spectrophotometer, Perkin-Elmer, MA, USA). Fluorescence spectra were determined using LS-55 fluorescence spectrophotometer (Perkin-Elmer, MA, USA). Upconversion luminescence spectra were obtained on the Ocean Insight spectrometer with Spetrasuite software (Florida, USA). The PA signal intensities were recorded by a 10 MHz, 10 mJ/cm², 384-element ring ultrasound array, and an optical parametric oscillator (OPO) (BB-OPO-NIR, Deyang-Tech, Zhejiang, China; pump laser, Nimma-900, Beamtech, Beijing, China) with 5–10 ns pulse duration and 10 Hz pulse repetition rate was used as the light source. The TEM images of UCNPs and UCNPs-DI were captured by a high-resolution 2100F field emission transmission electron microscope (JEOL, Japan) operating at a capture acceleration voltage of 200 kV. ZEN3690 zeta sizer (Malvern, USA) was used to measure the particle size. The Bio-Rad FTS 6000 spectrometer (Bio-Rad Company, Hercules, California, USA) was used to record the FT-IR spectrum in the form of KBr particles. Flow cytometry assay was performed on CytoFLEX (Beckman Coulter, USA). Fluorescence imaging of the cells was acquired on the Olympus Fluoview FV 3000 microscope (Olympus Imaging America, Japan).

For immunofluorescence microscopy studies, samples were prepared as described previously (47 (link), 48 (link)). Confocal images were acquired with a FluoView FV3000 microscope (Olympus) with a 40× oil immersion objective lens. Details are presented in Text S1.

H9c2 cells were seeded in chambers with coverslip (5 × 10⁴ cells per chamber), incubated for 48 h at 37 °C, 5% CO₂ and then treated with vehicle or PSELT-dansyl 30 nM for 15 min. Cells were washed with DPBS and fixed for 10 min with ice-cold methanol. Fixed cells were rinsed with DPBS two times and permeabilization phase was performed using 0.1 % Triton X-100 in DPBS for 30 min at RT. Permeabilized H9c2 cells were then washed with DPBS and blocked with 1% BSA in DPBS for 30 min at RT. For immunofluorescence analysis, cells were incubated with a primary antibody against laminin (diluted 1:100) overnight at 4 °C and then stained with a donkey anti-rabbit secondary antibody, Alexa Fluor 488 (diluted 1:400) for 1 h at RT. The cells were then washed twice with DPBS and incubated with propidium iodide (PI) for nuclei staining. Images were obtained using an Olympus Fluoview FV3000 microscope and the images were taken with FV31S-SW software. Quantification of fluorescence signals was carried out using CellSens Dimension software 1.7 (Count&Measure Full, Olympus Europa SE & Co., Hamburg, Germany).