Measurement of cellular 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction activity and lactate dehydrogenase release using MTT

3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction assay and lactate dehydrogenase (LDH) release assay have been widely used for evaluating cell viability in culture. MTT reduction assay measures the redox activity of living cells, while LDH assay measures the activity of L...

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Bibliographic Details
Published in:Neuroscience research 2000-12, Vol.38 (4), p.325-329
Main Authors: Abe, Kazuho, Matsuki, Norio
Format: Article
Language:English
Subjects:
3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT)
Animals
Animals, Newborn
Astrocyte
Astrocytes - drug effects
Astrocytes - metabolism
Biological Assay
Cell Survival - physiology
Cells, Cultured - drug effects
Cells, Cultured - metabolism
Colorimetry
Coloring Agents - pharmacology
Culture
Culture Media
Cytotoxicity assay
Cytotoxins - pharmacology
Formazans - metabolism
L-Lactate Dehydrogenase - metabolism
Lactate dehydrogenase
Neuron
Neurons - drug effects
Neurons - metabolism
Oxidation-Reduction
Rats
Rats, Wistar
Tetrazolium Salts - metabolism
Tetrazolium Salts - pharmacology
Thiazoles - pharmacology
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Summary:3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction assay and lactate dehydrogenase (LDH) release assay have been widely used for evaluating cell viability in culture. MTT reduction assay measures the redox activity of living cells, while LDH assay measures the activity of LDH released into the medium from dead cells. In this paper, we introduce a quick and simple method of measuring cellular MTT reduction and LDH release with the same dye, MTT. The substrate mixture for measuring LDH activity contained lactate, β-nicotinamide adenine dinucleotide, 1-methoxyphenazine methosulfate, MTT and Triton X-100. When the medium containing LDH was mixed with the substrates, MTT was converted into MTT formazan in proportion to LDH activity. This method was successfully applied for evaluating t-butyl hydroperoxide toxicity in cultured rat cortical astrocytes and glutamate toxicity in cultured rat hippocampal neurons. Our method is economical and convenient especially for measuring cellular MTT reduction and LDH release in the same culture.
ISSN:0168-0102
1872-8111
DOI:10.1016/S0168-0102(00)00188-7