Would like to measure oxidative stress in the brain tissue. I am thinking to measure lipid peroxidation using MDA levels. If anyone has any suggestions, to measure oxidative stress in the brain tissue lysate using other parameters, please let me know.
Note: -
Thank you,
Mayur Parmar
Hi Mayur, this should be a good one for your needs. best, George
Journal of Neuroscience Methods 144 (2005) 127–135
Lipid peroxidation measurement by thiobarbituric
acid assay in rat cerebellar slices
Yngo J. Garcia a, b, c, d, ∗, Antonio J. Rodr´ıguez-Malaver a, Nancy Pe˜naloza a
perform the assay in 1.5 ml tubes and finally read in 96 well plate , assay require a boiling step and u may not use the plate.. However alternative assays such as isoprostanes estimation which also indicate the lipid modifications can be done in standard 96 well plates using ELISA. attached here is working protocol for mda assay i routinely use. Do not use perfused of fixed brain slices (fixed in formaldehyde) it will not give MDA assay
Best
Dear Mayur. The kits for 96 plate of Oxford Biomedical Research is very economic (220 euro) and results is perfect, and protocol is easy for used. I have experience with this kit and results in liver, in brain.
Best regards.
Thanks Dr. Corcoran, Aditya and Kireew for your response. I will look into your suggestions.
Hi,
I would like to add some more information about the estimation of LPO in brain tissue. I am using this method in my Laboratory.
Method:
The malondialdehyde content, a measure of lipid peroxidation, was assayed in the form of thiobarbituric acid reactive substances by the method of Wills (1965). Briefly,
0.5 ml of postmitochondrial supernatant and 0.5 ml of Tris–HCl were incubated at 37°C for 2 h. After incubation, 1 ml of 10% trichloroacetic acid was added and centrifuged at 1,000 g for 10 min. To 1 ml of supernatant, 1 ml of 0.67%
thiobarbituric acid was added, and the tubes were kept in boiling water for 10 min. After cooling, 1 ml of double distilled water was added, and absorbance was measured at 532 nm. Thiobarbituric acid-reactive substances were quantified using an extinction coefficient of 1.56×105 M−1 cm−1 and expressed as nanomole of malondialdehyde per milligram protein. Tissue protein was estimated using the Biuret method, and the brain malondialdehyde content was expressed as nanomole of malondialdehyde (MDA) per milligram of protein (PMID: 21103863).
http://link.springer.com/article/10.1007%2Fs00213-010-2094-2
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