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Chloride Transport across the Lysosomal Membrane
Published in Bruno Gasnier, Michael X. Zhu, Ion and Molecule Transport in Lysosomes, 2020
Sonali Saha, Anja Blessing, Thomas J. Jentsch
Different fluorescence microscope setups can be used for intracellular Clˉ imaging. It is important to have a sensitive camera such as a CCD (charge-coupled device) camera. For imaging lysosomes, one should use an objective with high magnification (e.g. 63x or 100x) that is suited for fluorescence microscopy. Depending on the fluorophores, adequate filter sets must be selected. For MEQ, the set should allow excitation at 344 nm and enable fluorescence detection at 442 nm. TMR is excited at 550 nm while the emission peak is at 580 nm.
Basic Principles in Photomedicine and Photochemistry
Published in Henry W. Lim, Nicholas A. Soter, Clinical Photomedicine, 2018
The excited singlet state molecule formed upon absorption of a photon typically has a lifetime of less than 50 nsec (10–9 sec) and sometimes as short as a few psec (10–12 sec). Emission of fluorescence and internal conversion are photophysical processes that occur very rapidly and return the excited singlet state molecule to the ground state without producing a chemical change. Fluorescence is the emission of a photon of light by the excited singlet state. The wavelengths of fluorescence emission of a molecule are at longer wavelengths than the absorption wavelength of the molecule. Fluorescence has many applications in dermatology. For example, a Wood’s lamp, which emits mainly in the UVA range, is used to detect fluorescence associated with certain cutaneous changes such as porphyrins in the urine of patients with porphyria. Fluorescence from dyes such as fluorescein and rhodamine is used for fluorescence microscopy. The fluorescence of the crosslinking amino acid derivatives desmosine and pyridinoline is used to detect these compounds in elastin and collagen, respectively.
Ultraviolet Fluorescence Photography
Published in Adrian Davies, Digital Ultraviolet and Infrared Photography, 2017
When a substance fluoresces without the addition of a dye it is known as autofluorescent. Other materials can have dyes, or markers, added to them to produce fluorescence. These dyes are known as fluorophores (or fluorochromes) and are used, for example, in fluorescence microscopy to help identify different tissues. One particular fluorophore is the protein dye sodium fluorescein, used extensively in forensics, microscopy and ophthalmology.
Advances in imaging techniques to assess kidney fibrosis
Published in Renal Failure, 2023
Buchun Jiang, Fei Liu, Haidong Fu, Jianhua Mao
Recently, studies on fluorescence microscopy are limited to animal models. A study involving murine UUO models showed that SHG imaging found more collagen in the fibrotic kidney; the combination of SHG imaging and fluorescence lifetime imaging is a promising technique that can accurately assess the degree of kidney fibrosis [34]. Another study using HistoIndex’s Genesis200 platform in mice with UUO, which combines SHG imaging and two-photon excitation microscopy, showed that HistoIndex’s Genesis200 platform could sensitively detect the collagen deposition of the fibrotic kidney and is be a good supplement to traditional pathological examination [35]. Studies on fluorescence microscopy are still in the preclinical stage; however, they show the possibility of visual and dynamic observation of kidney fibrosis.
13-Acetoxysarcocrassolide induces apoptosis in human hepatocellular carcinoma cells through mitochondrial dysfunction and suppression of the PI3K/AKT/mTOR/p70S6K signalling pathway
Published in Pharmaceutical Biology, 2022
Chang-Min Hsu, Jen-Jie Lin, Jui-Hsin Su, Chih-I Liu
TUNEL/DAPI staining was performed using the In Situ Cell Death Detection Kit, Fluorescein (Roche Diagnostics, Indianapolis, IN, USA), and DAPI (Sigma-Aldrich, St. Louis, MO, USA). The cells were cultured on coverslips at 1 × 104 cells/well. After adhesion, the cells were treated with different concentrations of 13-acetoxysarcocrassolide for 24 h. Next, the culture medium was removed, and the cells were fixed with 4% paraformaldehyde for 20 min at room temperature and then washed three times with PBS before the cell membranes were permeabilized with 0.1% Triton-X100 for 2 min on ice. Subsequently, the cells were washed with cold PBS three times. Next, 30 μL of pre-chilled TUNEL staining solution and enzyme were mixed in a 1:19 ratio before adding them to the cells; this was incubated at 37 °C for 1 h. Cells were then washed three times with PBS and stained with 1 µL of DAPI diluted with 1 mL of PBS for 10 s. The samples were placed on a slide and observed under a fluorescence microscope.
Capillin protects against non-alcoholic steatohepatitis through suppressing NLRP3 inflammasome activation and oxidative stress
Published in Immunopharmacology and Immunotoxicology, 2021
Bin Li, Rui Wang, Lei Wang, Gucheng Zhang, Yang Zhang
The liver tissues with a thickness of 4 μm were embedded in paraffin with 10% formalin, and the paraffin-removed liver tissues were dyed with H&E. The staining effect was observed and analyzed by an optical microscope, and the pathological changes in HFD rat liver tissues were observed under 100- and 200-fold visual fields. Five fields of liver tissue were selected in each mouse and the pathological lesions were differentiated in all of them. The histopathological diagnosis was done independently by two pathologists. Histological activation index (HAI) was calculated to semi-quantitatively evaluate the severity of hepatic injury based on a prior study [26]. For NLRP3 detection in the liver of mice, the paraffin-embedded sections (4 μm) were dewaxed in dimethylbenzene, hydrated in gradient ethanol, and blocked in 5% goat serum. Afterward, the sections were incubated with anti-NLRP3 antibody (ab270449, Abcam) overnight at 4 °C. The nucleus was stained using 4′,6-diamidino-2-phenylindole (DAPI, Beyotime, Shanghai, China). The fluorescence signal was monitored by a fluorescence microscope.