Fluorescent probes 2′,7′-Dichlorodihydrofluoresce in diacetate (Solarbio, China) were used to detect intracellular ROS in cells. DCFH-DA as oxidative sensitive fluorescent probe allowed to assess specific intracellular ROS by being oxidised to fluorescent DCF (36 (link)).The loss of MMP was estimated by JC-1 kit according to manufacturer’s introduction (Beyotime, China). To perform assays, HaCaT cells (2 × 105 cells/well) were paved overnight. Then, the cells were pretreated with deucravacitinib or Deu@PEPS (0.5 μg/ml)and co-stimulated by TNF-α and IL-17A for 24 h. The cells were dyed with JC-1 dye or DCFH-DA probes (10 μM) in the sunblock and washed to remove remaining probes. The staining was tested using the fluorescence microscope.
2 7 dichlorodihydrofluoresce in diacetate
Manufactured by Solarbio
Sourced in China
2′,7′-Dichlorodihydrofluoresce in diacetate is a fluorogenic substrate used for the detection and measurement of esterase activity in biological samples. It is a non-fluorescent compound that becomes highly fluorescent upon hydrolysis by esterases, allowing for the quantification of enzyme activity.
Automatically generated - may contain errors
Lab products found in correlation
Jc 1 kit, by Beyotime (1 mentions)
Facscanto 2, by BD (1 mentions)
Facschorus software, by BD (1 mentions)
Fluorescence microscope, by Leica (1 mentions)
Cell counting kit 8 cck 8, by Beyotime (1 mentions)
Annexin 5 fitc apoptosis detection kit, by Solarbio (1 mentions)
Jem 1200ex, by JEOL (1 mentions)
Calcein pi cell viability cytotoxicity assay kit, by Beyotime (1 mentions)
Cytation 5, by Agilent Technologies (1 mentions)
Wst 8, by Beyotime (1 mentions)
Rpmi 1640 medium, by Thermo Fisher Scientific (1 mentions)
1 1 dioctadecyl 3 3 3 3 tetramethylindocarbocyanine perchlorate dii, by Beyotime (1 mentions)
Jc 1 mitochondrial membrane potential fluorescent probe, by Solarbio (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Solarbio (1 mentions)
Dulbecco s modified eagle medium dmem, by Thermo Fisher Scientific (1 mentions)
3 3 dioctadecyloxacarbocyanine perchlorate dio, by Beyotime (1 mentions)
Cell counting kit 8 cck 8, by Apexbio (1 mentions)
6 protocols using 2 7 dichlorodihydrofluoresce in diacetate
1
Intracellular ROS and Mitochondrial Membrane Potential Assays
2
ROS Quantification in Primary Rat Keratinocytes
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PRKs (2x105/ml) were incubated in 6-well plates with 2',7'-dichlorodihydrofluorescein diacetate (1.0 µM; Beijing Solarbio Science & Technology Co., Ltd.) at 37˚C for 15 min. Subsequently, PRKs were washed twice with PBS and analyzed by FCM (FACSCanto II; BD FACSChorus™ software, version: 1.0; BD Biosciences) to detect ROS using a 488-nm laser for excitation and a 535-nm laser for detection.
3
Assessing Oxidative Stress Markers in PC12 Cells
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PC12 cells were seeded in 24-well plates and allowed to adhere. When the groups finished processing for 1 day, the reactive oxygen species (ROS) fluorescent probe 2,7-dichlorodihydrofluorescein diacetate (Beijing Solarbio Science & Technology Co., Beijing, China, Cat# CA1410) was added to different groups of cells and incubated in the incubator for 30 minutes. Finally, cells were observed and photographed using a fluorescence microscope (Leica, Wetzlar, Germany).
Superoxide dismutase (SOD; Beijing Solarbio Science & Technology Co., Cat# BC0175), plasma glutathione peroxidase (GSH-Px), and malondialdehyde (MDA; Beijing Solarbio Science & Technology Co., Cat# BC0025) detection kits were used to detect SOD activity, GSH-Px levels, and MDA contents in accordance with the manufacturer’s instructions.
Superoxide dismutase (SOD; Beijing Solarbio Science & Technology Co., Cat# BC0175), plasma glutathione peroxidase (GSH-Px), and malondialdehyde (MDA; Beijing Solarbio Science & Technology Co., Cat# BC0025) detection kits were used to detect SOD activity, GSH-Px levels, and MDA contents in accordance with the manufacturer’s instructions.
4
Zinc-Based Nanomedicine for Cancer Therapy
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Zinc nitrate hexahydrate (Zn(NO3)2·6H2O, 99%) was obtained from Tianjin FuChen Chemical Reagent Co., Ltd. (Tianjin, China). 2-methylimidazole (C4H6N2, 98%) was purchased from TCI Development Co., Ltd. (Shanghai, China). Dihydromyricetin was secured from Shanghai Winherb Medical Technology Co., Ltd. (Shanghai, China) with purity >99%; PAN Seratech GmbH, Aidenbach, Germany, provided fetal bovine serum. Cell Counting Kit-8 (CCK-8) and Calcein/PI Cell Viability/Cytotoxicity Assay Kit were supplied by Shanghai Beyotime Biotechnology Co., Ltd. (Shanghai, China). Annexin V-FITC Apoptosis Detection Kit and 2′,7′-Dichlorodihydrofluorescein diacetate was purchased from Beijing Solarbio Science & Technology Co., Ltd. (Beijing, China). The antibodies were purchased from Abcam (Shanghai) trading Co., Ltd. (Shanghai, China). Absorbance and fluorescence intensity were detected using the TECAN spark multifunctional microplate reader. Fluorescence images were taken with a Nikon Tclipse Ts2R fluorescence inverted microscope. The apoptosis assay was performed with the BD FACSaira II flow cytometer. The apoptotic morphology was photographed using a JEM-1200EX (JEOL, Tokyo, Japan) transmission electron microscope.
5
Antioxidant Activity Assays for imDCs
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To assess superoxide dismutase (SOD) activity, imDCs at a density of 1×106 cells/mL were washed twice with cold PBS, homogenized in ice-cold PBS, subjected to a total SOD assay with WST-8 (Beyotime, China) at 37°C for 30 min, and then had their absorbance read by a microplate spectrophotometer (Cytation5, Biotek) at 450 nm. To evaluate glutathione peroxidase (GPx) activity, imDCs at a density of 1×106 cells/mL were homogenized, subjected to a cellular glutathione peroxidase assay (Beyotime, China), and then had their absorbance read by a microplate spectrophotometer at 340 nm. To measure ROS levels, imDCs at a density of 1×106 cells/mL were washed twice with PBS, resuspended in PBS, incubated with 2′, 7′-dichlorodihydro-fluorescein diacetate (Solarbio, China) for 20 min at 37oC, and then had their activity determined by flow cytometry assay [34 (link)]. To determine glutathione (GSH) content, imDCs at a density of 1×106 cells/mL were washed with PBS, repeatedly frozen and thawed with a protein removal reagent, and then had their absorbance read by a microplate spectrophotometer [35 (link)].
6
Celastrol-loaded Nanoparticles for Cancer Therapy
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Celastrol was bought from Chengdu Zhibiaohuachun Biotechnology (Chengdu, China); betulinic acid, cholesterol and coumarin-6, from J&K Scientific (Beijing, China); 1.2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy (polyethylene glycol)-2000] -folic acid and soybean phospholipid (SPC), from Xi’an Ruixi Biotechnology (Xi’an, China); fetal bovine serum (FBS), calf serum, RPMI-1640 medium, and Dulbecco’s modified Eagle medium (DMEM), from GIBCO (Thermo Fisher Scientific, Carlsbad, California, USA); 3.3′-dioctadecyloxacarbocyanine perchlorate (DiO) and 1.1’-dioctadecyl-3,3,3’,3’-tetramethylindocarbocyanine perchlorate (DiI), from Beyotime Biotechnology; and DAPI, from Solarbio Life Sciences (Beijing, China).
The Cell Counting Kit-8 (CCK-8) was obtained from APExBIO Technology (Houston, TX, USA). 2’,7’-Dichlorodihydrofluorescein diacetate and JC-1 mitochondrial membrane potential fluorescent probe was obtained from Solarbio (Beijing, China). Antibodies were obtained from Abcam (Cambridge, UK) against α-SMA (catalog no. ab124964), Bcl-2 (ab196495) or collagen I (ab260043). Antibody against P-glycoprotein 1 (catalog no. AF5185) was obtained from Affinity Biosciences (Jiangsu, China).
The Cell Counting Kit-8 (CCK-8) was obtained from APExBIO Technology (Houston, TX, USA). 2’,7’-Dichlorodihydrofluorescein diacetate and JC-1 mitochondrial membrane potential fluorescent probe was obtained from Solarbio (Beijing, China). Antibodies were obtained from Abcam (Cambridge, UK) against α-SMA (catalog no. ab124964), Bcl-2 (ab196495) or collagen I (ab260043). Antibody against P-glycoprotein 1 (catalog no. AF5185) was obtained from Affinity Biosciences (Jiangsu, China).
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