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Reduction of MTT by glutathione S-transferase
J L York1, L C Maddox, P Zimniak
1University of Arkansas for Medical Sciences, Little Rock 72205, USA.
Biotechniques
|October 30, 1998
Summary
Glutathione S-transferase (GST) enzymes can directly reduce the MTT assay reagent, leading to inaccurate cell viability measurements. This finding necessitates caution when interpreting results, particularly in studies involving GST modulation.
Area of Science:
- Biochemistry
- Cell Biology
- Toxicology
Context:
- The MTT assay is a standard method for assessing cell viability and proliferation.
- This assay relies on the reduction of a tetrazolium salt (MTT) by mitochondrial dehydrogenases in living cells.
- Elevated formazan product levels can indicate increased metabolic activity or interference with the assay.
Purpose:
- To investigate the potential interference of glutathione S-transferase (GST) enzymes with the MTT assay.
- To determine if GST enzymes can directly catalyze the reduction of MTT.
- To evaluate the implications of GST activity on the interpretation of cell survival and proliferation data.
Summary:
- The study found that glutathione S-transferase (GST) enzymes, particularly GST P1-1, can directly reduce the tetrazolium salt 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) in vitro.
- Overexpression of mouse GST P1-1 resulted in significantly high background levels of the formazan product in cell cultures.
- Purified GST enzymes confirmed their ability to catalyze MTT reduction, independent of cellular mitochondrial activity.
Impact:
- Cytotoxicity assays utilizing the MTT assay require careful interpretation, especially when investigating compounds that affect GST expression or activity.
- The findings highlight a potential confounding factor in studies relying on MTT for cell viability assessment.
- Researchers should consider alternative assays or controls when GST modulation is a variable.