Lactate dehydrogenase (LDH) release, superoxide dismutase (SOD) activity, and malondialdehyde content were assessed via colorimetric assays, as measures of oxidative stress. Three days after irradiation the supernatant of the culture medium was collected, and LDH release was measured using a commercially available LDH assay kit (Nanjing Jiancheng, Nanjing, China) in accordance with the manufacturer’s instructions. SOD activity and malondialdehyde content in cells were assessed via SOD assay kits and malondialdehyde assay kits (Nanjing Jiancheng).
Malondialdehyde assay kit
Manufactured by Nanjing Jiancheng
Sourced in China
The Malondialdehyde assay kit is a laboratory tool used to measure the level of malondialdehyde, a biomarker for oxidative stress. The kit provides reagents and protocols for colorimetric or fluorometric quantification of malondialdehyde in various sample types.
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Lab products found in correlation
Superoxide dismutase assay kit, by Nanjing Jiancheng (6 mentions)
Reduced glutathione assay kit, by Nanjing Jiancheng (5 mentions)
Total superoxide dismutase assay kit, by Nanjing Jiancheng (3 mentions)
Peroxidase assay kit, by Nanjing Jiancheng (3 mentions)
Glutathione peroxidase assay kit, by Nanjing Jiancheng (2 mentions)
Total protein quantitative assay kit, by Nanjing Jiancheng (2 mentions)
Total antioxidant capacity assay kit, by Nanjing Jiancheng (2 mentions)
Catalase assay kit, by Nanjing Jiancheng (2 mentions)
Ldh assay kit, by Nanjing Jiancheng (1 mentions)
Sod assay kit, by Nanjing Jiancheng (1 mentions)
Eclipse ti microscope, by Nikon (1 mentions)
Mnsod assay kit with wst 8, by Beyotime (1 mentions)
Hexadecyltrimethylammonium bromide, by Merck Group (1 mentions)
Protein assay kit, by Bio-Rad (1 mentions)
Enhanced chemiluminescence reagent, by Santa Cruz Biotechnology (1 mentions)
Horseradish peroxidase linked secondary antibody, by Santa Cruz Biotechnology (1 mentions)
O dianisidine dihydrochloride, by Merck Group (1 mentions)
Hematoxylin and eosin, by Merck Group (1 mentions)
Probucol, by Merck Group (1 mentions)
Anti β actin antibody, by Merck Group (1 mentions)
31 protocols using malondialdehyde assay kit
1
Oxidative Stress Assessment Protocol
2
ROS Assay and Oxidative Stress Quantification
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After treatment and washing with PBS 3 times, cells were incubated with reactive oxygen species (ROS) assay mixture according to the protocol of the reactive oxygen species (ROS) assay kit. Five random fields were observed and imaged per slide using a Nikon ECLIPSE Ti microscope (Nikon, Tokyo, Japan). The levels of malondialdehyde (MDA) were determined by using commercially available total superoxide dismutase and malondialdehyde assay kits (Nanjing Jiancheng Bioengineering Institute, China) according to the manufacturer's protocol. The MnSOD assay kit with WST-8 (Beyotime Institute of Biotechnology, Shanghai, China) was used to measure SOD2 enzymatic activity according to the manufacturer's instructions.
3
Antioxidant and Anti-Inflammatory Assay
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Probucol, anti-β-actin antibody, hexadecyltrimethylammonium bromide, o-dianisidine dihydrochloride, and hematoxylin and eosin were purchased from Sigma Chemical Company (St. Louis, MO, USA). Protein assay kit was obtained from Bio-Rad Laboratories (Hercules, CA, USA). The anti-E-selectin antibody, horseradish peroxidase-linked secondary antibody, and enhanced chemiluminescence reagent were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Malondialdehyde assay kit was obtained from Nanjing Jiancheng Institute of Bioengineering (Nanjing City, China). All other reagents were of the highest grade commercially available.
4
Lung Tissue Biochemical Analyses
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Lung tissues from each group were homogenized in pre-cooled PBS, and were frozen in liquid nitrogen and thawed three times. Following centrifugation at 10,000 × g for 10 min at 4°C, the supernatant was collected, in order to determine the concentrations of hydroxyproline, MDA and GSH, and the activity of SOD. HO-1 activity was evaluated by determining the amount of bilirubin, as previously described (19 (link)). All measurements were performed using commercial kits (Hydroxyproline assay kit, Malondialdehyde assay kit, Glutathione Peroxidase assay kit, Total Superoxide Dismutase assay kit, and Total Bilirubin kit) obtained from Nanjing Jiancheng Bioengineering Institute (Jiangsu, China) according to the manufacturer's manual.
5
Oxidative Stress Biomarkers in Liver
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The ROS was detected by Reactive Oxygen Species Fluorogenic Probe (BestBio; BB470513) and was observed with confocal microscopy. The frozen liver tissue (100 mg) was homogenized in 0.9 ml PBS. The lysate was centrifuged and the supernatant was collected. Hepatic MDA content, GSH content, iron content, TRF content was detected using the Malondialdehyde assay kit (Jiancheng, A003‐1‐2), Reduced glutathione assay kit (Jiancheng, A006‐2‐1), tissue iron assay kit (Jiancheng, A039‐2‐1), and Transferrin Assay Kit (Jiancheng, E028‐1‐1) following the manufacturer's instructions. The values for the levels of GSH in the liver tissues were measured using a microplate reader at the absorption wavelengths of 405nm. The values for the levels of MDA, iron, TRF in the liver tissues were measured using a spectrophotometer at the absorption wavelengths of 532, 520, and 340 nm.
6
Serum Biomarkers for Transplant Monitoring
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Venous blood of recipient pigs from the 2 groups were sampled at postoperative days 1–5. Blood samples were centrifuged at 3000 rpm for 10 min, and supernatants were harvested for determining serological levels of tumor necrosis factor-a (TNF-α), interleukin-1 (IL-1), interleukin-6 (IL-6) and malondialdehyde (MDA) were detected using an enzyme-linked immunosorbent assay (ELISA) according to the manufacturer’s protocol (Pig TNF-α ELISA kit Ab100756, IL-1 Beta Pig ELISA kit Ab10075, IL-6 Pig ELISA kit Ab100755, Abcam, Cambridge, UK, and Malondialdehyde assay kit (A003-1 Nanjing Jiancheng Bioengineering Institute, Nanjing, China). Levels obtained are expressed as either pg/mL or mmol/mL.
7
Serum Biochemical Profile Quantification
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Total protein (TP), albumin (ALB), total cholesterol (TC), triglyceride (TG), nonesterified fatty acid (NEFA), total antioxidant capacity (TAOC), and malondialdehyde (MDA) in serum or muscle (n = 8 per treatment) were tested respectively by total protein quantitative assay kit (A045-2-2), albumin assay kit (A028-2-1), total cholesterol assay kit (A111-1-1), triglyceride assay kit (A110-1-1), nonesterified free fatty acids assay kit (A042-2-1), total antioxidant capacity assay kit (A015-1-2 ), and malondialdehyde assay kit (A003-1-2 ) purchased from Nanjing Jiancheng Bioengineering Institute (Nanjing, China) in accordance with the manufacturer's instructions.
8
Oxidative Stress Biomarkers Measurement
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The levels of malondialdehyde, glutathione, and iron were measured by Malondialdehyde Assay Kit (TBA method) (A003-1-2, Jiancheng Biotech, Nanjing, China), Reduced Glutathione Assay Kit (A006-2-1, Jiancheng Biotech) and Iron Assay Kit (A009-2-1, Jiancheng Biotech) according to the manufacturer’s instructions, and the corresponding absorbance was detected by a microplate reader to calculate the concentration (the determination wavelength of malondialdehyde, glutathione and irons are 532 nm, 420 nm and 520 nm, respectively). The formula for calculating malondialdehyde concentration is the concentration of malondialdehyde (nmol/mgprot) = (OD sample − OD Background)/(OD Standard − OD Background) × 10 (nmol/ml)/concentration of sample protein (mgprot/ml). The formula for calculating glutathione concentration is the concentration of glutathione (μmol/gprot) = (OD sample − OD Background)/(OD Standard − OD Background) × 20 (μmol/l)/concentration of sample protein (gprot/l).
9
Serum SOD and MDA Measurement
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A Superoxide Dismutase Assay Kit (A001-1; Nanjing Jiancheng Bioengineering Institute, China) was selected for the serum SOD measurements. A Malondialdehyde assay kit (A003-1; Nanjing Jiancheng Bioengineering Institute, China) was chosen for the serum MDA measurements. The assays were conducted according to the manufacturer’s instructions.
10
Antioxidant Markers Quantification in Brain Tissue
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Firstly, the tissue was rapidly homogenized in ice-cold PBS (9 times of tissue weight) using a homogenizer. After centrifugation at 4500 g for 15 min at 4 °C, the supernatant was collected to determine the protein concentration using a protein assay kit (Beyotime, China). For measuring SOD, the Superoxide Dismutase Detection Kit (Nanjing Jiancheng Bioengineering Institute, Nanjing, China) was used. According to the manufacturer's instruction [26 (link)], the assay was carried out. GSH serves as the body's most significant non-enzymatic antioxidant and scavenges free radicals. As directed by the manufacturer's instruction, we employed a Reduced glutathione assay kit (Nanjing Jiancheng Bioengineering Institute, Nanjing, China) to ascertain the level of GSH present in brain tissue. GSH may form a yellow molecule when it reacts with dithiodinitrobenzoic acid (DTNB), which enables colorimetric quantitative measurement of GSH content at 405 nm. MDA is a marker of lipid peroxidation. In accordance with the manufacturer's instruction from the Malondialdehyde assay kit (Nanjing Jiancheng Bioengineering Institute, Nanjing, China), we employed thiobarbituric acid (TBA) to measure the MDA concentration [26 (link), 27 (link)]. Finally, a colorimetric test at 532 nm was performed on the supernatant in each tube.
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