GSH and GSSG assay kit (Beyotime Biotech, China), reactive oxygen species (ROS) assay kit, Maleic Dialdehyde (MDA) assay kit and iNOS assay kit (Jiancheng Biotech, China) were used to determine the oxidative stress level in liver or cell. All the results were normalized to protein concentrations for animal studies or normalized to cell numbers for in vitro experiments.
Inos assay kit
Manufactured by Nanjing Jiancheng
Sourced in China
The INOS assay kit is a laboratory equipment designed to detect and quantify the levels of nitric oxide synthase (iNOS) in biological samples. It provides a reliable and standardized method for researchers to analyze iNOS activity, which is an important enzyme involved in various physiological and pathological processes.
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Lab products found in correlation
Gsh and gssg assay kit, by Beyotime (1 mentions)
Ros assay kit, by Nanjing Jiancheng (1 mentions)
Anti f4 80 fitc, by Thermo Fisher Scientific (1 mentions)
Anti cd11b fitc, by Thermo Fisher Scientific (1 mentions)
Adenosine triphosphate (atp), by Merck Group (1 mentions)
Lipopolysaccharides (lps), by Merck Group (1 mentions)
Phorbol myristate acetate (pma), by Merck Group (1 mentions)
Dextran sulfate sodium (dss), by MP Biomedicals (1 mentions)
Exfect transfection reagent, by Vazyme (1 mentions)
Mpo activity assay kit, by Nanjing Jiancheng (1 mentions)
Murine elisa kits, by Neobioscience (1 mentions)
7600 110 autoanalyzer, by Hitachi (1 mentions)
Mitochondrial extraction kit, by Solarbio (1 mentions)
Malondialdehyde mda assay kit, by Solarbio (1 mentions)
Atp assay kit, by Nanjing Jiancheng (1 mentions)
Chamq universal sybr qpcr master mix, by Vazyme (1 mentions)
Hiscript 3 rt supermix for qpcr gdna wiper, by Vazyme (1 mentions)
Sod assay kit, by Nanjing Jiancheng (1 mentions)
Gsh px assay kit, by Nanjing Jiancheng (1 mentions)
Cat assay kit, by Nanjing Jiancheng (1 mentions)
7 protocols using inos assay kit
1
Oxidative Stress Biomarker Assays
2
Oroxylin A Purification and Inflammatory Assays
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Oroxylin A was isolated from the root of Scutellaria baicalensis as previously described [31 (link)]. Samples containing oroxylin A at a minimum of 99% purity were used for the experiments unless otherwise indicated. LPS, PMA and ATP were purchased from Sigma Aldrich (St. Louis, MO, USA). DSS (molecular weight 36–50 kDa) was obtained from MP Biomedicals Inc. (Irvine, CA, USA). Antibodies against p-IκBα, IκBα and p65 were purchased from Cell Signaling Technology (Danvers, MA, USA). Primary antibodies against lamin A, β-actin, IL-1β and bay11-7082 were purchased from Bioworld Technology Inc. (CA, USA). Exfect Transfection reagent was purchased from Vazyme (Nanjing, China). NLRP3 and caspase-1 antibodies were from Abcam Technology Inc (MA, USA). FITC-anti-CD11b and FITC-anti-F4/80+ were purchased from eBioscience (San Diego, CA, USA). ELISA kits for murine IL-1β and human IL-1β were purchased from Boster Biotech Co. Ltd. (Wuhan, China). MPO activity assay kit and iNOS Assay Kit were purchased from Nanjing Jiancheng Bioengineering Institute (Nanjing, China). Immunohistochemistry kit was purchased from KeyGEN Biotech Inc. (Nanjing, China).
3
Measuring Inflammatory Cytokines and iNOS
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The inducible nitric oxide synthase (iNOS) activity was assayed using an iNOS assay kit (Jiancheng, Nanjing, China). The TNF-α, IL-12, and IL-10 levels in supernatants were detected using Murine ELISA kits (Neobioscience, China) according to the manufacturer's instructions.
4
Myocardial Infarction in Mice
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C57BL/6 male mice were purchased from an experimental animal company. XTL continuous zoom stereomicroscope (Shenzhen Ruiwode Life Technology Company, Shenzhen, China) and MicroVent 1 small animal ventilator (Pittsfield, USA) were used in the microsurgical operations. Hitachi 7600-110 autoanalyzer was used for biochemical analyses. Chloral hydrate and isoflurane were obtained from Sun Chemical Technology (Shanghai, China). In addition, microsurgical instruments, endotracheal intubations, and disposable intravenous catheters (22 G) were used in this study. The iNOS assay kit was obtained from Nanjing Jiancheng Bioengineering Institute (Nanjing, China).
Thirty C57BL/6 male mice were randomly divided into three groups (n=10 mice/group): a sham-operation group (sham), an AMI operation group (AMI), and a nitrite pretreatment+AMI operation group (N+AMI). Before the AMI operation, mice in the N+AMI group were pretreated with sodium nitrite in drinking water (50 mg/L in double distilled water) for 7 days according to previous studies (11 (link),12 (link)). Mice were housed in controlled temperature, humidity, and 12-h light–dark cycle with free access to chow and water. Mice studies were approved by the Ningbo University Institutional Animal Care and Use Committee.
Thirty C57BL/6 male mice were randomly divided into three groups (n=10 mice/group): a sham-operation group (sham), an AMI operation group (AMI), and a nitrite pretreatment+AMI operation group (N+AMI). Before the AMI operation, mice in the N+AMI group were pretreated with sodium nitrite in drinking water (50 mg/L in double distilled water) for 7 days according to previous studies (11 (link),12 (link)). Mice were housed in controlled temperature, humidity, and 12-h light–dark cycle with free access to chow and water. Mice studies were approved by the Ningbo University Institutional Animal Care and Use Committee.
5
Intracellular iNOS Activity Measurement
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For the detection of intracellular iNOS activity, RAW264.7 macrophages at the concentration of 3 × 106 were washed with ice-cold PBS (pH 7.2) three times after removal of the cell culture medium. Then, the macrophages were suspended in 1 ml of ice-cold PBS and centrifuged at 1000 × g for 5 min. For the next, the macrophages were re-suspended in 50 μl of ice-cold cell lysis buffer (20 mM Tris–HCl pH 7.5, 150 mM NaCl, 1 mM EDTA, 1 mM EGTA, 1 mM glycerol, 1% Triton X-100, 1 mM DTT, 1 μg/ml Leupeptin, 1 mM PMSF). After 20 min incubation on ice, the resulting solution was vortexed for 20 min and centrifuged at 15,000 × g for 10 min at 4°C in a centrifuge (Eppendorf, Germany). After the determination of protein concentration in the detergent-soluble fraction, the aliquots were stored at −80°C for further experiments. iNOS assay kit purchased from Nanjing Jiancheng Bioengineering Institute, China was utilized to measure the iNOS activity following the product instruction.
6
Biochemical Assays for Avian Viral Infection
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ATP assay kit, superoxide dismutase (SOD ) assay kit, GSH-PX assay kit, catalase (CAT ) assay kit, and inducible nitric oxide synthase (iNOS ) assay kit were obtained from Nanjing Jiancheng Bioengineering Institute (Nanjing, China). Mitochondrial extraction kit, cytochrome c oxidase (COX ) assay kit, malondialdehyde (MDA ) assay kit, and succinate dehydrogenase (SDH ) assay kit were purchased from Beijing Solarbio Science & Technology Co., Ltd. (Beijing, China). HiScript III RT SuperMix for qPCR (+gDNA wiper) and ChamQ Universal SYBR qPCR Master Mix were purchased from Vazyme (Jiangsu, China). Dulbecco's Modified Eagle's Medium (DMEM , Hyclone, Logan, UT,) supplemented with penicillin 100 IU/mL, streptomycin 100 IU/mL, and 10% fetal bovine serum was used as a nutritive medium. The concentration of fetal bovine serum in DMEM was replaced to 1% as maintenance medium (MM). D-Hank's solution was used for washing the embryo tissue fragments and cells. Baicalin (85%) was purchased from Shanghai Jieli Biotechnology Co., Ltd. (Shanghai, China). The DHAV-1 (strain LQ2) used in the challenge experiments was supplied by the Shandong Institute of Poultry (Shandong, China) and was diluted into 50 tissue culture infectious dose 50 with MM.
7
Enzyme Inhibition Assay Protocol
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The inhibition of LOX activity was determined based on a method previously described [35 (link)], and linoleic acid was used as the substrate.
The inhibition of COX-2 activity was determined using a cyclooxygenase inhibition screening kit (Beyotime Biotechnology, Shanghai, China).
The inhibition of iNOS activity was determined using the iNOS assay kit (Nanjing Jiancheng Bioengineering Institute, Nanjing, China).
The inhibition of COX-2 activity was determined using a cyclooxygenase inhibition screening kit (Beyotime Biotechnology, Shanghai, China).
The inhibition of iNOS activity was determined using the iNOS assay kit (Nanjing Jiancheng Bioengineering Institute, Nanjing, China).
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