Oxidative stress measurements were performed on the dissected hippocampi and cortices of four rats from each group sacrificed at four time points (5 min, 30 min, 24 h, and 3 days) after KA or pilocarpine treatment respectively (S0033, Beyotime Institute of Biotechnology, China). We dissected out the hippocampus and cortex. These different regions were soaked separately in cell staining buffer (0.01 M phosphate-buffered saline [PBS], Shanghai Novland Co., Ltd., China). Filtration was used to prepare a single-cell suspension, which was incubated with a ROS marker, 2′,7′-dichlorofluorescin diacetate (DCF-DA, S0033, Beyotime Institute of Biotechnology, China) at 37 °C for 60 min. Tissues were then washed three times with PBS. A fluorescence microplate reader (Thermo, USA) was used to measure excitation at 488 nm and emission at 525 nm.
S0033
Manufactured by Beyotime
Sourced in China
The S0033 is a laboratory equipment product. It serves as a primary function without further interpretation or extrapolation.
Automatically generated - may contain errors
Lab products found in correlation
Vert a1, by Zeiss (3 mentions)
S0063, by Beyotime (3 mentions)
Synergy h1, by Agilent Technologies (2 mentions)
Mitosox, by Thermo Fisher Scientific (2 mentions)
Mitosox red mitochondrial superoxide indicator, by Thermo Fisher Scientific (2 mentions)
Fv1000, by Olympus (1 mentions)
Eclipse ts2r, by Nikon (1 mentions)
Cover glass bottom dish, by SPL Life Sciences (1 mentions)
Rhod 2 am, by Thermo Fisher Scientific (1 mentions)
Fluo 4 am, by Thermo Fisher Scientific (1 mentions)
Laser confocal microscope, by Olympus (1 mentions)
Flow cytometry, by BD (1 mentions)
Ckx41, by Olympus (1 mentions)
Mito tracker green, by Beyotime (1 mentions)
Facscaliburtm, by BD (1 mentions)
Evos fl, by Thermo Fisher Scientific (1 mentions)
S0101, by Beyotime (1 mentions)
S0056, by Beyotime (1 mentions)
Ldh c0017, by Beyotime (1 mentions)
Microplate reader, by Bio-Rad (1 mentions)
52 protocols using s0033
1
Oxidative Stress Measurement in Rat Brain Regions
2
ROS Quantification using H2DCF-DA
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ROS levels were determined using an active oxygen detection kit (s0033, Biyuntian Biotechnology Ltd., Wuhan, China). Briefly, cells were incubated with 5 μM H2DCF-DA (chloromethyl-2′, 7′-dichlorofluorescein diacetate) for 30 min at 37 °C to load the fluorescent dye.
3
Oxidative Stress Measurement Protocols
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The oxidative stress was evaluated by determining the levels of malondialdehyde (MDA) and reactive oxygen species (ROS). According to the instructions, the MDA detection kit for lipid peroxidation (S0131, Biyuntian, China) and the active oxygen detection kit (S0033, Biyuntian, China) were used for detection, respectively.
4
ROS Detection in AVIC Cells
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The level of cellular ROS was measured using a ROS detection kit (S0033, Beyotime, China). Briefly, AVICs were seeded in a 96-well plate (4000 cells/well), cultured with PM and three concentrations of metformin for 72 h. Moreover, 100 μl of 10 μM DCFH-DA reagent was added to each well and incubated at 37 ℃ for 30 min. Cells cultured in normal medium were used as a negative control. Excessive dyes were removed using M199 medium without FBS. A fluorescence microplate reader was used to measure the fluorescence intensity value, with excitation wavelength of 488 nm and emission wavelength of 525 nm.
5
Measuring Intracellular ROS in HepG2 Cells
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Production of intracellular reactive oxygen species (ROS) in HepG2 cells was determined based upon the oxidation of DCFH-DA (S0033, Beyotime Company, Shanghai, China). Fluorescence emission at 525 nm following excitation at 488 nm was measured using a microplate reader. The cellular fluorescence intensity was expressed as the fold change relative to the level observed in the control cells. The cellular fluorescence intensity was expressed as the fold change relative to the level observed in the control cells.
6
Measuring Mitochondrial Oxidative Activity
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Mitochondrial reactive oxygen species and total oxidative activity were analyzed using the mitochondrial fluorescent probe MitoSox (M36008; Thermo Fisher, USA) and a fluorescent intracellular reactive oxygen species kit (S0033; Beyotime Biotech, China), respectively, according to a previously reported methodology [24 (link)]. Images were acquired using a confocal laser-scanning microscope (FV 1000; Olympus, Japan).
7
Hypoxia-Induced Oxidative Stress Evaluation
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Raw264.7 cells were cultured under normal oxygen or hypoxic conditions for 8 h. Oxidative stress levels were measured by detecting reactive oxygen species (ROS; S0033; Beyotime, China), according to the manufacturer’s instructions. The fluorescence signal was recorded using 520- and 590-nm lasers and detected using a fluorescence microscope (ECLIPSE TS2R; Nikon, Japan).
8
Intracellular ROS Detection Assay
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To detect intracellular reactive oxygen species (ROS), we used an ROS assay kit (S0033, Beyotime). All the procedures were performed according to the manufacturer’s instructions. Briefly, after washing twice with sterile PBS, cells were stained with 10 μM DCFDA at 37 ℃ for 20 min in the dark. Then, the cells were washed with basal culture medium three times. The images were captured using a fuorescence microscope (ZEISS Vert. A1).
9
Intracellular ROS Detection Assay
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To detect intracellular reactive oxygen species (ROS), we used an ROS assay kit (S0033, Beyotime). All the procedures were performed according to the manufacturer’s instructions. Briefly, after washing twice with sterile PBS, cells were stained with 10 μM DCFDA at 37 ℃ for 20 min in the dark. Then, the cells were washed with basal culture medium three times. The images were captured using a fuorescence microscope (ZEISS Vert. A1).
10
Calcium and ROS Imaging Assays
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The cells were seeded in a cover glass‐bottom dish (SPL Life Sciences Co., Ltd., Korea). Calcium mobilization analysis was performed using Fluo‐4 AM (acetoxymethyl) ester (Invitrogen, San Diego, CA, USA) or Rhod‐2 AM (Invitrogen, San Diego, CA, USA), as described in the recent report.[12 ] Intracellular ROS were measured by flow cytometry using a cell‐based ROS assay kit (S0033; Beyotime Biotechnology, Beijing, China) and MitoSOX (M36008, Invitrogen, USA). Details of the procedure are available in Appendix A1 in the Supporting Information.
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