ATP content in rat myocardial tissues was inspected using the ATP assay kit (#S0026; Beyotime, Beijing). Rat myocardial tissue homogenates in lysis buffer were centrifuged. The supernatants were mixed with ATP detection solution and kept at room temperature for 5 min. The levels of ATP were determined by measuring the relative light unit (RLU) with a luminometer, using the standard curve obtained from 0.01, 0.03, 0.1, 0.3, 1, 3 and 10 μM ATP samples as a reference. MDA levels were analyzed by the lipid peroxidation MDA assay kit (Colorimetric/Fluorometric) (#ab118970; Abcam) according to the manufacturer’s instructions. SOD activity was measured using the SOD assay kit (#19160; Sigma-Aldrich), as specified by the manufacturer (21 (link), 22 (link)).
Lipid peroxidation mda colorimetric fluorometric assay kit
Manufactured by Abcam
Sourced in United States
The Lipid Peroxidation (MDA) Colorimetric/Fluorometric Assay Kit is a laboratory tool used to measure the level of malondialdehyde (MDA), a marker for lipid peroxidation. The kit provides a colorimetric or fluorometric method to quantify MDA in biological samples.
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Lab products found in correlation
Glutathione assay kit, by Cayman Chemical (3 mentions)
Sod assay kit wst, by Dojindo Laboratories (2 mentions)
Sod assay kit, by Merck Group (1 mentions)
Atp assay kit, by Beyotime (1 mentions)
Hydrogen peroxide assay kit, by Abcam (1 mentions)
Superoxide dismutase sod colorimetric activity kit, by Thermo Fisher Scientific (1 mentions)
Glutathione peroxidase assay kit, by Cayman Chemical (1 mentions)
Hydrogen peroxide colorimetric detection kit, by Enzo Life Sciences (1 mentions)
Aas800, by PerkinElmer (1 mentions)
Synergy htx microplate reader, by Agilent Technologies (1 mentions)
Catalase activity colorimetric fluorometric assay kit, by Abcam (1 mentions)
Gssh gsh quantification kit, by Dojindo Laboratories (1 mentions)
Protein carbonyl elisa kit, by Enzo Life Sciences (1 mentions)
8 protocols using lipid peroxidation mda colorimetric fluorometric assay kit
1
Myocardial ATP, MDA, and SOD Analysis
2
Oxidative Stress Biomarkers Quantification
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On day 42, the hind paw tissues were homogenized with 100 μL ice-cold tissue RIPA lysis buffer. The homogenate was incubated at 4 °C for 30 min and centrifuged at 12,000× g at 4 °C for 20 min. The supernatant was obtained as the total protein extract. Total protein concentration was measured with the bicinchoninic acid (BCA) assay kit.
The level of malondialdehyde (MDA) was determined with the Lipid Peroxidation (MDA) Assay Kit (Colorimetric/Fluorometric) (Cat# ab118970, Abcam, Cambridge, MA, USA) according to the manufacturer’s instructions. Superoxide Dismutase (SOD) and glutathione peroxidase (GSH-Px) activities were determined with the Superoxide Dismutase (SOD) Colorimetric Activity Kit (Cat# EIASODC, Thermo Fisher Scientific, Waltham, MA, USA) and glutathione peroxidase ELISA Kit (Cat#11352, Cusabio Biotech Co., Ltd., Wuhan, China), all according to manufacturer’s instructions. The amount of hydrogen peroxide (H2O2) was estimated using the Hydrogen Peroxide Assay Kit (ab102500, Abcam, Cambridge, MA, USA) according to the manufacturer’s instructions.
The level of malondialdehyde (MDA) was determined with the Lipid Peroxidation (MDA) Assay Kit (Colorimetric/Fluorometric) (Cat# ab118970, Abcam, Cambridge, MA, USA) according to the manufacturer’s instructions. Superoxide Dismutase (SOD) and glutathione peroxidase (GSH-Px) activities were determined with the Superoxide Dismutase (SOD) Colorimetric Activity Kit (Cat# EIASODC, Thermo Fisher Scientific, Waltham, MA, USA) and glutathione peroxidase ELISA Kit (Cat#11352, Cusabio Biotech Co., Ltd., Wuhan, China), all according to manufacturer’s instructions. The amount of hydrogen peroxide (H2O2) was estimated using the Hydrogen Peroxide Assay Kit (ab102500, Abcam, Cambridge, MA, USA) according to the manufacturer’s instructions.
3
Oxidative Stress Biomarkers Measurement
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The plasma total glutathione (T-GSH) level and the erythrocyte GSH and GSSG levels were measured using a glutathione assay kit (Cayman Chem, Ann Arbor, MI). Plasma and erythrocyte GR activities were measured using a GSH reductase assay kit (Cayman Chem), and plasma and erythrocyte GPx activities were measured using a glutathione peroxidase assay kit (Cayman Chem) according to the manufacturer's instructions. The erythrocyte glutamate-cysteine ligase, catalytic subunit (GCLC) level was measured using a GCLC ELISA kit (MyBioSource, Inc, San Diego, CA) [28] .
As a parameter to estimate systemic oxidative stresses, we measured the plasma malondialdehyde (MDA) level using a Lipid Peroxidation (MDA) Assay Kit (Colorimetric/Fluorometric; Abcam, Cambridge, MA) [29] . To evaluate the ability of the plasma MDA level to estimate oxidative stresses, the erythrocyte H 2 O 2 level was measured in rats using a hydrogen peroxide colorimetric detection kit (Enzo Life Science, Farmingdale, NY), and then its correlation with the plasma MDA level was analyzed.
In addition, the plasma selenium level was measured by electrothermal atomic absorption spectrometry using a spectrometer equipped with an Atomic Absorption Spectrophotometer (AAS 800; PerkinElmer Inc, San Jose, CA) [30] .
As a parameter to estimate systemic oxidative stresses, we measured the plasma malondialdehyde (MDA) level using a Lipid Peroxidation (MDA) Assay Kit (Colorimetric/Fluorometric; Abcam, Cambridge, MA) [29] . To evaluate the ability of the plasma MDA level to estimate oxidative stresses, the erythrocyte H 2 O 2 level was measured in rats using a hydrogen peroxide colorimetric detection kit (Enzo Life Science, Farmingdale, NY), and then its correlation with the plasma MDA level was analyzed.
In addition, the plasma selenium level was measured by electrothermal atomic absorption spectrometry using a spectrometer equipped with an Atomic Absorption Spectrophotometer (AAS 800; PerkinElmer Inc, San Jose, CA) [30] .
4
Quantifying Lipid Peroxidation via MDA
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The level of MDA was determined using the Lipid Peroxidation (MDA) Colorimetric/Fluorometric Assay kit (BioVision, Inc.) by ELISA. The skin tissue was homogenized with MDA Lysis Buffer. It was subsequently centrifuged at 13,000 x g for 10 min. A total of 200 µl of supernatant was then place in a 1.5 ml microcentrifuge tube. Thiobarbituric acid (TBA; 600 µl) was then added to each well and incubated at 95°C for 60 min. The sample was placed in the ice for 10 min and thawed to the room temperature (20-25°C). A total of 200 µl was taken from 800 µl reaction mixture to a 96-well plate for analysis. The Synergy HTX microplate reader (BioTek Instruments, Inc.) was used to measure the absorbance at 532 nm. The level of MDA was calculated according to the manufacturer's instructions.
5
Lipid Peroxidation and SOD Assessment
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The lipid peroxidation was determined by measuring the levels of malondialdehyde (MDA) using a Lipid Peroxidation (MDA) Colorimetric/Fluorometric Assay Kit (BioVision), according to the manufacturer’s instructions. Total superoxide dismutase (SOD) activities were assessed using a SOD assay kit – WST (Dojindo, Kumamoto, Japan) following the manufacturer’s instructions.
6
Evaluation of Levetiracetam's Effects on Hormones and Oxidative Stress
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LEV (Keppra®, UCB Pharma) tablets were used for experiments. FSH, LH, and testosterone levels in serum were determined by ELISA kits from Cusabio Biotech Co. Ltd., Hubei, P. R. C. GSH levels were measured with Cayman® Glutathione Assay Kit, MI, USA. Also, MDA, CAT levels and SOD activity were determined by BioVision® Lipid Peroxidation (MDA) Colorimetric/Fluorometric Assay Kit, CA, USA; BioVision® Catalase Activity Colorimetric/Fluorometric Assay Kit, CA, USA; and BioVision® Superoxide Dismutase (SOD) Activity Assay Kit, CA, USA; respectively.
7
Oxidative Stress Biomarker Assessment
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The liver tissue was homogenized on ice in 300 μL of malondialdehyde (MDA) lysis buffer (with 3 μL of BHT (100×) and then centrifuged (13,000 × g, 10 min) to remove insoluble material. Lipid peroxidation of the plasma was determined with a Lipid Peroxidation (MDA) Colorimetric/Fluorometric Assay Kit (BioVision; CA, U.S.A.). Glutathione peroxidase (GSH-Px) activity in plasma was measured using the Cayman (MI, U.S.) assay kit and superoxide dismutase (SOD) in the liver tissue was determined with a SOD Assay Kit-WST (Dojindo; MD, U.S.A.). GSH of liver tissues were determined with GSSH/GSH Quantification Kit (Dojindo; MD, U.S.A.) according to the manufacturer’s instructions.
8
Oxidative Stress Biomarkers in Liver
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Reduced (GSH) and oxidized (GSSG) glutathione were measured in liver tissue extracts using the Glutathione Assay Kit purchased from Cayman Chemicals (Montigny-le-Bretonneux, France).
Hepatic carbonylated proteins were also assessed as valuable markers of oxidative stress [28] (link).
Protein carbonyl quantification was carried out using the Protein Carbonyl Elisa Kit purchased from Enzo Life Sciences (Villeurbanne, France). Lipid peroxidation was assessed using the Lipid Peroxidation (MDA) Colorimetric/Fluorometric Assay Kit (K739) from BioVision. Protein expression of hepatic cytochrome P450 2E1 (CYP2E1) and heat shock cognate 70 (HSC70) (used as loading control) was assessed by western blot analysis as previously described [15] (link), using respectively the antibodies from Oxford Biomedical Research (Oxford, MI) and Santa Cruz Biotechnology (Dallas, TX). The dilution used for the antibodies was 1:1000 for CYP2E1 and 1:500 for HSC70.
Hepatic carbonylated proteins were also assessed as valuable markers of oxidative stress [28] (link).
Protein carbonyl quantification was carried out using the Protein Carbonyl Elisa Kit purchased from Enzo Life Sciences (Villeurbanne, France). Lipid peroxidation was assessed using the Lipid Peroxidation (MDA) Colorimetric/Fluorometric Assay Kit (K739) from BioVision. Protein expression of hepatic cytochrome P450 2E1 (CYP2E1) and heat shock cognate 70 (HSC70) (used as loading control) was assessed by western blot analysis as previously described [15] (link), using respectively the antibodies from Oxford Biomedical Research (Oxford, MI) and Santa Cruz Biotechnology (Dallas, TX). The dilution used for the antibodies was 1:1000 for CYP2E1 and 1:500 for HSC70.
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