Dear all, I have followed the below protocol to assess the MDA (malondialdehyde) in the mouse optic nerve. As I know the reaction between MDA and thiobarbituric acid (TBA) should yield pink color which absorbed max at 532nm. But I did not get any pink color, the reaction mixture was changed to yellow instead. Do any of you have the same problem or do you have any idea about that? Thanks in advance.
My protocol is as follow:
1- Homogenized in PBS
2- Keep the homogenate at 37 ͦC for 1 hour.
3- 25 µl of the homogenate were mixed with 15 µl of BHT (0.5 mg/ml ethanol) and 30 µl of TCA 25%. The mixture was incubated at 100 ͦ C for 10 min.
4- Centrifuged at 2000g for 10 min, 4 degrees of centigrade.
5- Taken supernatant.
6- The sample were mixed with 50 µl TBA % 0.33 at 95 ͦ C for 1 hour.
The colour of solution might turn into yellow after incubation by adding BHT.
Also the OD value should be taken at 530nm.
https://www.researchgate.net/publication/325056897_Elevated_serum_levels_of_malondialdehyde_and_cortisol_are_associated_with_major_depressive_disorder_A_case-control_study?_sg%5B0%5D=7h9hg8ZXhc1YBy_yPdCfrITxExi7tSZYtG3slAXMYiuPmhIybPp0NC72TvdPfxOThsPgEx-579kxIXcQT6dBMJukQBXRH9CJltEELmqr.CZjpDEY1_-UzySjQH4IHjtYrne8a5V8cS9o8RnNcLUvLP-JZDvHW8ShZ0HdQVZIuCxag5oxKUYa7jnVSpBPTFg
- Badges
- Science topic
- Similar topics
- Biological Science
- Anatomy
- Tissue