Lipid Peroxidation (MDA + HNE) Assay Kit

Lipid peroxidation is a well-known example of oxidative damage to cell membranes, lipoproteins and other lipid-containing structures. Peroxidative modification of unsaturated phospholipids, glycolipids and cholesterol can occur in various reactions. They are characterized by i) free radical species such as oxal radicals, peroxyl radicals, and hydroxyl radicals derived from the iron-mediated reduction of hydrogen peroxide or ii) non-radical species such as singlet oxygen, ozone, and peroxynitrite superoxide produced by reaction with nitric oxide.

Malondialdehyde (MDA) and 4-hydroxyalkenals are important toxic byproducts of lipid peroxidation. So, the measurement of the amount of such aldehydes corresponds to an index of lipid peroxidation in vitro and in vivo. 4-Hydroxynonenal (4-HNE)-6 is a major product of the peroxidative decomposition of polyunsaturated fatty acids (PUFAs). It has cytotoxic, hepatotoxic, mutagenic and genotoxic properties. Furthermore, increased levels of HNE were found in the plasma and various organs under oxidative stress conditions. In fact, MDA is in many instances the most abundant individual aldehyde resulting from lipid peroxidation. In vitro MDA can transform proteins, DNA, RNA and many other biomolecules.

Bioquochem’s LPO Assay Kit measures MDA and HNE concentrations as an index of lipid peroxidation. First, the acid-catalyzed attack on the 3-position of the indole ring initiates the reactions between the indole and the aldehyde (MDA and HNE). As a result, this reaction gives a diindolylkane (chromophore) with maximum absorption in the region of 580–620 nm.

In our assay an indole (reagent A) reacts quickly with MDA and HNE in acidic medium, generating a chromophore (C) with a high molar extinction coefficient at a maximum absorption wavelength of 586 nm.

Product Overview

The Lipid Peroxidation (MDA) Assay Kit (Colorimetric/Fluorometric) (ab118970) provides a convenient tool for the sensitive detection of malondialdehyde (MDA).

In the lipid peroxidation assay protocol, the MDA in the sample reacts with thiobarbituric acid (TBA) to generate the MDA-TBA adduct. MDA-TBA adduct can be easily quantified colorimetrically (OD = 532 nm) or fluorometrically (Ex/EM = 532/553 nm). This assay detects MDA levels of at least 1 nmol/well colorimetrically and 0.1 nmol/well fluorometrically.

The MDA assay is also known as the TBARS assay.

Lipid Peroxidation Assay Protocol Summary:
– Add TBA solution to samples and standards, incubate at 95 °C for 60 min, cool in an ice bath for 10 min
– Transfer to wells of microplate
– Microplate Reader
For higher sensitivity, precipitate with n-butanol, centrifuge, dry and re-suspend pellet before analysis.

Chinese protocol is available. See Protocol section below.

For an alternative MDA assay, try MDA assay ab233471, without the heating steps required in the TBARS assay.

notes
Lipid peroxidation refers to the oxidative degradation of lipids. In this process free radicals take electrons from lipids (usually in the cell membrane), resulting in cell damage. Quantification of lipid peroxidation is essential for assessing oxidative stress. Lipid peroxidation forms reactive aldehydes as natural bi-products such as malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE). Malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE) are often used as markers of lipid peroxidation and to assay for oxidative damage/oxidative stress.

related product

Review the Oxidative Stress Marker and Assay Guide, or the Complete Metabolic Assay Guide, to learn more about assaying for metabolites, metabolic enzymes, mitochondrial function, and oxidative stress, and also in live cells using your plate reader. How to test metabolic function.

Also refer to the popular 4-HNE assay kit ab238538 as an alternative marker of lipid peroxidation and oxidative stress.

How have other researchers used the Lipid Peroxidation Assay Kit ab118970?

The MDA/TBARs assay kit has been used in publications in a variety of sample types, including:
– Human: Serum 1, Hippocampal Primary Cell Extracts 2, A375 Cultured Cell Lysates 3, Plasma and Platelet Samples 4
– Mouse: neuronal cell lysates 5 , cardiac tissue extracts 6 , plasma 7 , cell extracts 8
– Rat: Hippocampal tissue extract 9 , Cardiomyocyte extract of cultured cells 10 , Lung lysates 11
– Pig: Serum 12

References: 1 – Shen J et al. 2018, 2 – Wang Q et al. 2019, 3 – Luo M et al. 2018, 4 – Mustafa AG et al. 2018, 5 – Murphy K et al. 2018, 6 – Guan F et al. 2019, 7 – Costa CRC et al. 2018, 8 – Eleftheriadis T et al. 2019, 9 – Malekian et al. 2019, 10 – Zhou Z et al. 2018, 11 – Lee L et al. 2018, 12 – Lee SE and Kang KS 2019

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