H2o2 assay kit

Manufactured by Nanjing Jiancheng

Sourced in China

The H2O2 assay kit is a laboratory equipment used to quantitatively measure the concentration of hydrogen peroxide (H2O2) in various sample types. It provides a simple and accurate method for the detection and analysis of H2O2 levels.

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H2O2 kit (10 citations) Hydrogen Peroxide (H2O2) Content Assay Kit (2 citations)

34 protocols using h2o2 assay kit

Intestinal Antioxidant Evaluation

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Approximately 100 mg of frozen jejunum and colon tissues was removed quickly and homogenized with ice-cold physiologic saline (1 : 9, w/v) and then centrifuged at 2,000 g for 20 min at 4°C. The intestinal supernatants were used for further analysis. Plasma and intestinal antioxidant indicators, including catalase (CAT), superoxide dismutase (SOD), glutathione (GSH), and glutathione peroxidase (GSH-Px), as well as malondialdehyde (MDA) were analyzed by ELISA assay kits from Jiangsu Meimian Institute (Mei mian, Yancheng, China). The total antioxidant capacity (T-AOC) and H₂O₂ assay kits were purchased from Nanjing Jiancheng Bioengineering Institute (Nanjing, China). The test for each index was carried out according to the instructions of the kits. The absorbance values were read on a Multiscan Spectrum Spectrophotometer (Tecan, Infinite M200 Pro, Switzerland). The jejunal and colonic mucous antioxidant parameters were normalized to the total protein concentration (mg/L) quantified by the Pierce BCA Protein Assay Kit (CoWin Biosciences, Suzhou, China).

Wang K., Kong X., Azad M.A., Zhu Q., Xiong L., Zheng Y., Hu Z., Yin Y, & He Q. (2021). Maternal Probiotic or Synbiotic Supplementation Modulates Jejunal and Colonic Antioxidant Capacity, Mitochondrial Function, and Microbial Abundance in Bama Mini-piglets. Oxidative Medicine and Cellular Longevity, 2021, 6618874.

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Yeast-Induced Oxidative Defense in Fruits

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The disinfected fruits were inoculated on their circumference using an inoculating needle (5.0 mm). For inoculations, a 10 μL aliquot of the yeast suspension at a concentration of 1.0 × 10⁸ cells/mL was dropped onto each prick. After air drying, the fruits were stored in enclosed plastic trays to maintain a high humidity (approximately 95 %). The plastic cases were maintained at 28 °C for the indicated periods. To measure the elicitation effect, the tissue surrounding each wound of fruit was collected at hour 0, 12, 24, 36, 48, 60 after treatment, and immediately immersed in liquid nitrogen and stored at −80 °C until use. A 10-g sample (fresh weight; FW) of the exocarp was ground into a powder in liquid nitrogen.
The concentration of H₂O₂ was assayed using H₂O₂ assay kits (Nanjing Jiancheng Bioengineering Institute, China) according to the manufacturer’s instructions. The enzyme activities were determined by a Shimadzu UV-1800 spectrophotometer (Shimadzu, Japan). The activities of chitinase and β-1,3-glucanase were measured as described previously [11 (link)]. The lignin content was quantified using the method described by Syros et al. [66 (link)].

Liu P., Chen K., Li G., Yang X, & Long C.A. (2016). Comparative transcriptional profiling of orange fruit in response to the biocontrol yeast Kloeckera apiculata and its active compounds. BMC Genomics, 17, 17.

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Quantifying Hepatic Hydrogen Peroxide

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H₂O₂ accumulation was determined using a H₂O₂ Assay Kit (Nanjing Jiancheng Bioengineering Institute, China). Snap-frozen liver tissue was homogenized with saline. H₂O₂ concentration was then measured following the manufacturer’s instructions.

Peng M., Shao M., Dong H., Han X., Hao M., Yang Q., Lyu Q., Tang D., Shen Z., Wang K., Kuang H, & Cao G. (2023). Nanodrug rescues liver fibrosis via synergistic therapy with H2O2 depletion and Saikosaponin b1 sustained release. Communications Biology, 6, 184.

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Spectrophotometric Quantification of H2O2, NO, and PAL

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The levels of H₂O₂ and NO in plant tissues were determined using the spectrophotometric method with an H₂O₂ assay kit (Nanjing Jiancheng). The PAL enzyme activity in leaves was determined with a PAL detection kit (Nanjing Jiancheng).

Mo S., Zhang Y., Wang X., Yang J., Sun Z., Zhang D., Chen B., Wang G., Ke H., Liu Z., Meng C., Li Z., Wu L., Zhang G., Duan H, & Ma Z. (2021). Cotton GhSSI2 isoforms from the stearoyl acyl carrier protein fatty acid desaturase family regulate Verticillium wilt resistance. Molecular Plant Pathology, 22(9), 1041-1056.

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Measuring Antioxidant Capacity in Plant Tissues

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A certain weight of leaf or shoot sample was crushed in a prechilled pestle and crushed with a mortar in an appropriate volume of prechilled extraction buffer. After uniform homogenization and centrifugation at 4,000 g for 15 min at 4°C, the supernatant was collected and used for biochemical analyses.
Antioxidative capabilities were estimated by quantifying peroxidation products and antioxidase activity levels. The malondialdehyde (MDA) content was determined using a Plant MDA Assay Kit (TBA method; A003-3-1, Nanjing Jiancheng, Nanjing, China). The hydrogen peroxide (H 2 O 2 ) level was determined using an H 2 O 2 Assay Kit (H 2 O 2 reacted with titanium salt and is then quantified at OD 415 ; A064-1-1, Nanjing Jiancheng), and the superoxide anion (O 2 -) level was determined using an O 2 -Assay Kit (sulfanilamide method; R30342, Shanghai Yuanye, Shanghai, China). The activity levels of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) were determined using the Total SOD (WST-1 method; A001-1-2, Nanjing Jiancheng), POD (H 2 O 2 catalysis method; A084-3-1, Nanjing Jiancheng) and CAT (H 2 O 2 catalysis method; A007-1-1, Nanjing Jiancheng) Assay Kits, respectively, following the manufacturer's instructions. All the leaf and shoot samples were measured three times in parallel.

Sun H., Wu S., & Wu L. (2021). White oak (Quercus fabri Hance) regenerated stump sprouts show few senescence symptoms during 40 years’ growth in a natural forest.

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Histochemical Analysis of H2O2 and MDA

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Histochemical staining of H₂O₂ was performed using DAB solution (Sigma‒Aldrich) at a working concentration of 1 mg/ml. C. lanceolata PEMs were infiltrated with DAB solution and incubated in the dark for 1.5 h at room temperature. The stained PEMs were removed and made into slides, which were observed and photographed using an inverted microscope (Leica, DMI4000) and a confocal microscope (Carl Zeiss, LSM 800). The H₂O₂ content was determined using an H₂O₂ assay kit (Jiancheng, China), and the absorbance was measured at 415 nm using an enzyme labelling instrument with three biological replicates. The MDA content was determined using an MDA assay kit (Jiancheng, China), and the absorbance was measured at 532 nm using an enzyme labelling instrument with three biological replicates.

Hao Z., Shi J., Wu H., Yan Y., Xing K., Zheng R., Shi J, & Chen J. (2023). Phytosulfokine contributes to suspension culture of Cunninghamia lanceolata through its impact on redox homeostasis. BMC Plant Biology, 23, 480.

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Evaluating Oxidative Changes in Injured Mycelia

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To evaluate changes in ROS, injured mycelia of the CK and KA groups were collected at 15, 30, and 60 min after damage and immediately frozen at -80°C. The samples were ground with liquid nitrogen. Inhibition of superoxide anion (O^2-) activity was performed as described by the Inhibition and produced O^2- assay kit (Nanjing Jiancheng Bioengineering Institute, Nanjing, China). In this reaction system, the amount of O^2- inhibited by 1 g of mycelial protein at 37°C for 40 min was related to the amount of O^2- inhibited by 1 mg of Vc, as monitored at 550 nm, and this value was consider as one unit of O₂^--inhibiting activity. Hydrogen peroxide (H₂O₂) detection was performed as described by the H₂O₂ assay kit (Nanjing Jiancheng Bioengineering Institute, Nanjing, China), as monitored at 405 nm; hydroxy free radical (-OH) detection was performed as described by the -OH assay kit (Nanjing Jiancheng Bioengineering Institute, Nanjing, China), as monitored at 550 nm. The O₂^--inhibiting activity and H₂O₂ and -OH concentrations were calculated using the mycelial protein concentration.

Zhang J., Chen H., Chen M., Wang H., Wang Q., Song X., Hao H, & Feng Z. (2017). Kojic acid-mediated damage responses induce mycelial regeneration in the basidiomycete Hypsizygus marmoreus. PLoS ONE, 12(11), e0187351.

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Determination of O2- and H2O2 Contents

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The content of

O_{2}^{-}

and H₂O₂ was measured using

O_{2}^{-}

Content Test Kit and H₂O₂ Assay Kit (Jian Cheng Bioengineering Institute, Nanjing, China), respectively; 0.5 g of sample was added with 4.5 ml physiological saline and ground into homogenate. After incubation on ice for 2 h, the mixture was centrifuged at 8,000 g for 30 min, and the supernatant was used. The corresponding reagent was added according to the manufacturer's instructions, and

O_{2}^{-}

and H₂O₂ contents were determined by measuring the OD at 550 nm and 405 nm, respectively. Both

O_{2}^{-}

and H₂O₂ contents were expressed as mmol/g Prot.

Han Y., Yang R., Wang Q., Wang B, & Prusky D. (2022). Sodium silicate promotes wound healing by inducing the deposition of suberin polyphenolic and lignin in potato tubers. Frontiers in Plant Science, 13, 942022.

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Antioxidant Capacity Analysis of Plant Extracts

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A certain weight of leaf or shoot sample was crushed in a prechilled pestle and crushed with a mortar in an appropriate volume of prechilled extraction buffer. After uniform homogenization and centrifugation at 4,000 g for 15 min at 4°C, the supernatant was collected and used for biochemical analyses.
Antioxidative capabilities were estimated by quantifying peroxidation products and antioxidase activity levels.
The malondialdehyde (MDA) content was determined using a Plant MDA Assay Kit (TBA method; A003-3-1, Nanjing Jiancheng, Nanjing, China). The hydrogen peroxide (H 2 O 2 ) level was determined using an H 2 O 2 Assay Kit (H 2 O 2 reacted with titanium salt and is then quanti ed at OD 415 ; A064-1-1, Nanjing Jiancheng), and the superoxide anion (O 2 -) level was determined using an O 2 -Assay Kit (sulfanilamide method; R30342, Shanghai Yuanye, Shanghai, China). The activity levels of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) were determined using the Total SOD (WST-1 method; A001-1-2, Nanjing Jiancheng), POD (H 2 O 2 catalysis method; A084-3-1, Nanjing Jiancheng) and CAT (H 2 O 2 catalysis method; A007-1-1, Nanjing Jiancheng) Assay Kits, respectively, following the manufacturer's instructions. All the leaf and shoot samples were measured three times in parallel.

Sun H., Wu S., & Wu L. (2020). White Oak (Quercus fabri Hance) regenerated stump sprouts show few symptom of senescence during 40 years’ growth in the natural forest.

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Quantifying Reactive Oxygen Species

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The H₂O₂ content was determined using a H₂O₂ Assay Kit (Nanjing Jiancheng Bioengineering Institute, China).
The O₂^•- content was measured using Plant SOA Elisa Kit (Huijia Biotechnology Institute, Xiamen, China).

Zhang L., Hu T., Amombo E., Wang G., Xie Y, & Fu J. (2017). The Alleviation of Heat Damage to Photosystem II and Enzymatic Antioxidants by Exogenous Spermidine in Tall Fescue. Frontiers in Plant Science, 8, 1747.

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