After the indicated treatment, the cells were collected by trypsin digestion and washed with PBS. Then, 3 × 105 cells were acquired from each sample and mixed with 60 µL protein detergent S solvent from a GSH measurement kit (S0052, Beyotime). After sufficient vortex, the cells were lysed by three cycles of rapid freeze–thaw using liquid nitrogen and a 37 °C water bath. The samples were rested on ice for 5 min, centrifuged at 10,000× g for 10 min at 4 °C, and the supernatants were taken for the following GSH measurement by using a commercial kit (S0052, Beyotime), according to the manufacturer’s instructions.
S0052
Manufactured by Beyotime
Sourced in China
The S0052 is a laboratory equipment item. It is designed for specific laboratory applications. The core function of this product is to perform measurements and analyses in a controlled laboratory environment.
Automatically generated - may contain errors
Lab products found in correlation
S0131, by Beyotime (3 mentions)
S0109, by Beyotime (2 mentions)
S0103, by Beyotime (1 mentions)
Varioskan, by Thermo Fisher Scientific (1 mentions)
P0013, by Beyotime (1 mentions)
Synergy h1 microplate reader, by Synergy Software (1 mentions)
Kac1211, by Thermo Fisher Scientific (1 mentions)
Kac1261, by Thermo Fisher Scientific (1 mentions)
Khc3011, by Thermo Fisher Scientific (1 mentions)
Gsh px, by Beyotime (1 mentions)
S0053, by Beyotime (1 mentions)
9 protocols using s0052
1
Quantification of Cellular Glutathione
2
Oxidative Stress Markers Analysis
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Oxidative stress related markers of superoxide dismutase (SOD) (S0103, Beyotime, Shanghai, China) and glutathione (GSH) (S0052, Beyotime, Shanghai, China) were detected. The testis tissue or cell lysate was extracted and measured according to manufacturer’s instruction. Optical density (OD) values were read using microplate reader (Varioskan, Thermo Scientific, Shanghai, China) and analyzed by ImageJ software.
3
Measurement of Glutathione and Malondialdehyde in Brain
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Mice were perfused transcardially with PBS to remove blood. The brainstem samples were collected and were homogenized in ice-cold lysis buffer containing 20 mM Tris (pH 7.5), 150 mM NaCl, 1% Triton X-100, and protease inhibitor mixture. After 10 min of centrifugation at 10,000×g (4 °C), the contents of GSH and MDA in the collected supernatant were measured with commercial kits (GSH, S0052; MDA, S0131, Beyotime, Shanghai, China) according to the protocol provided by the manufacturer and previous studies.42–45 (link) The rationale for the measurement of GSH based on that GSH reacts with chromogenic substrate DTNB to produce yellow products and the amount of GSH can be detected by measuring absorption at A412. While, MDA can react with thiobarbituric acid (TBA) in high temperature and acidic environment to form red MDA-TBA adduct. The MDA-TBA adduct has the maximum absorption at 535nm, which can be detected by colorimetric method.
4
Lipid Peroxidation, Antioxidants, and Iron Levels
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A lipid peroxidation (malondialdehyde (MDA)) assay kit (S0131S, Beyotime, China), a total glutathione assay kit (S0052, Beyotime, China), and an iron assay kit (BC1735, Solarbio Science & Technology Co., Ltd. Beijing, China) were used to evaluate the lipid peroxidation product MDA and reduced glutathione (GSH) levels and ferrous iron (Fe2+) concentrations following the manufacturers’ instructions.
5
Oxidative Stress Biomarkers in PC12 Cells
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The activity of superoxide dismutase (SOD), GSH and MDA content was determined using the corresponding assay kits (S0109, S0052, S0131; Beyotime Biotechnology). PC12 cells (1 × 105 cells·mL−1) were seeded in six‐well plates and then 100 μm H2O2 was added, with treatment for 6 h. Next, the cells were cultured with or without 50 μm IRA‐3 and resveratrol for 24 h. Then PC12 cells were washed with PBS, and the cells were collected and lysed with cell lysis buffer for western blotting and immunoprecipitation (P0013; Beyotime Biotechnology) treatment. After centrifugation at 14 000 g for 5 min at 4 °C, the supernatant fractions were collected. The assays were conducted according to the kit instructions using a Microplate Reader.
6
Glutathione Assay in Sperm Samples
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Total glutathione (GSH) levels were evaluated following the protocol described by Zhu et al. [40 (link)] through a total glutathione assay kit (S0052, Beyotime Institute of Biotechnology, Shanghai, China). Briefly, sperm samples were centrifuged at 1000× g for 5 min at room temperature, re-suspended with a three-fold volume of Modena solution, and centrifuged at 1000× g for 10 min after cytolysis by three cycles of rapid cooling in liquid nitrogen and thawing at 37.8 °C. The supernatant was transferred into a 96-well plate to evaluate the absorption at 412 nm.
7
Measuring Oocyte Glutathione Levels
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Glutathione content was determined using a total glutathione assay kit (S0052, Beyotime Institute of Biotechnology, Haimen, China). Briefly, the matured oocytes from four groups (n ≥ 40 for each group, repeated 3 times) were pipetted repeatedly until lysis was completed. Glutathione contents of the oocytes from four groups were measured as described previously [18 (link)].
8
Quantifying Cytokines and Oxidative Stress
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The concentration of secreted protein was assessed by commercial ELISA kits depending on the method proposed by the manufacturer. For the detection of inflammation factors, tumor necrosis factor-α (TNF-α, KHC3011, Invitrogen, CA, USA), interleukin-6 (IL-6, KAC1261, Invitrogen, CA, USA), interleukin-1β (IL-1β, KAC1211, Invitrogen, CA USA), and oxidative stress indexes, malonic dialdehyde (MDA, S0131, Beyotime, Shanghai, China), superoxide dismutase (SOD, S0109, Beyotime, Shanghai, China) and glutathione peroxidase (GSH-px, S0052, Beyotime, Shanghai, China) in culture medium, the culture supernatant of HMCs after different treatments were collected and centrifuged to remove the cell debris. The clear supernatant was used for ELISA based on manufacturer’s instructions. Subsequently, the optical density (OD) value was detected using a Synergy H1 microplate reader (Winooski, Vermont, USA). The standard curve was created based on the concentration and OD value of the standard molecule provided in the kit. Finally, the concentration of cytokines and oxidative molecules were calculated based on the standard curve.
9
Glutathione Quantification in Intestinal Tissue
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