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Microtiter plate assay for superoxide dismutase using MTT reduction by superoxide
M Madesh1, K A Balasubramanian
1Wellcome Trust Research Laboratory, Department of Gastrointestinal Sciences, Christian Medical College Hospital, Vellore, India.
Indian Journal of Biochemistry & Biophysics
|November 6, 1998
Summary
A new colorimetric assay using microtiter plates accurately measures superoxide dismutase (SOD) activity. This simple method is ideal for high-throughput screening of SOD enzyme levels.
Area of Science:
- Biochemistry
- Enzymology
- Analytical Chemistry
Background:
- Superoxide dismutase (SOD) is a crucial antioxidant enzyme.
- Accurate measurement of SOD activity is vital for research and diagnostics.
- Existing SOD assays can be complex or require specialized equipment.
Purpose of the Study:
- To develop a simple, rapid, and sensitive microtiter plate-based colorimetric assay for superoxide dismutase (SOD) activity.
- To validate the assay's performance against established methods.
- To provide a cost-effective tool for high-throughput SOD activity measurements.
Main Methods:
- Superoxide generation via pyrogallol autoxidation.
- Inhibition of MTT dye reduction by SOD.
- Colorimetric measurement of formazan product at 570 nm after DMSO solubilization.
Main Results:
- The assay demonstrated sensitivity and stability of the colorimetric endpoint.
- Results correlated well with other known methods for purified Cu,ZnSOD and rat tissue samples.
- The microtiter plate format proved suitable for analyzing numerous samples efficiently.
Conclusions:
- A simple, inexpensive, and sensitive microtiter plate assay for SOD activity has been established.
- This method is suitable for routine laboratory use and large-scale screening.
- The assay provides a valuable tool for studying oxidative stress and antioxidant defense mechanisms.