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A rapid, quantitative microplate assay for NAD-linked D-mannitol dehydrogenase
Letters in Applied Microbiology
|October 1, 1990
Summary
A new microplate assay rapidly measures D-mannitol dehydrogenase activity using MTT reduction. This method is effective for bacterial extracts, permeabilized cells, and enzyme purification, correlating well with traditional techniques.
Area of Science:
- Biochemistry
- Enzymology
- Microbiology
Background:
- D-mannitol dehydrogenase is an important NAD-linked enzyme.
- Accurate and rapid enzyme activity measurement is crucial for research and purification.
Purpose of the Study:
- To describe a novel 96-well microtitre plate assay for NAD-linked D-mannitol dehydrogenase.
- To validate the assay's utility with bacterial extracts and during enzyme purification.
Main Methods:
- Enzyme activity measured via the reduction of MTT by NADH.
- Absorbance read at 620 nm using a microplate reader.
- Assay applied to crude bacterial extracts, permeabilized cells, and column eluates.
Main Results:
- The assay provides rapid measurement of D-mannitol dehydrogenase activity.
- Absorbance increase is directly proportional to enzyme concentration.
- Excellent correlation observed between microplate assay and conventional spectrophotometric methods.
Conclusions:
- The described microplate assay is a rapid, reliable, and efficient method for quantifying D-mannitol dehydrogenase.
- This assay is suitable for various biological samples and enzyme purification processes.