Evaluation of MTT assay for measurement of emodin-induced cytotoxicity
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Published on: November 10, 2013
Xiangfei Wang1, Junbo Ge, Keqiang Wang
1Shanghai Institute of Cardiovascular Diseases, Zhongshan Hospital, Fudan University, Shanghai, People's Republic of China.
Abstract:
The specificity and sensitivity of the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) tetrazolium assay can be influenced by certain factors, such as colored substances and cell volume. When the MTT assay is applied to measure cytotoxicity induced by emodin, its accuracy might be affected by emodin itself. Vascular smooth muscle cells were cultured in M199 medium. The optical density of emodin or formazan was measured by spectrophotometry. Emodin has a different absorption spectrum in different solvents. The solvents containing water induced a red shift of the absorption curve of emodin, which increased the overlap of the absorption curves of emodin and formazan. Formazan was formed from the MTT tetrazolium salt by emodin in a dose-dependent manner, which was partially suppressed by serum. Cytotoxicity was induced by emodin in a time- and dose-dependent manner in a modified MTT assay. The data suggest that emodin can alter the accuracy of the MTT assay but that a modified MTT assay is still valuable in measuring emodin-induced cytotoxicity.
Insights
Emodin interferes with the MTT assay
Area of Science:
- Biochemistry
- Cell Biology
- Pharmacology
Background:
- The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay is a common method for measuring cell viability and cytotoxicity.
- Factors like colored substances and cell volume can affect MTT assay accuracy.
- Emodin, a compound with potential therapeutic applications, may interfere with MTT assay measurements due to its properties.
Purpose of the Study:
- To investigate the impact of emodin on the accuracy of the MTT assay.
- To evaluate the reliability of a modified MTT assay for measuring emodin-induced cytotoxicity in vascular smooth muscle cells.
Main Methods:
- Culturing vascular smooth muscle cells in M199 medium.
- Measuring optical density of emodin and formazan using spectrophotometry.
- Assessing emodin's absorption spectrum in various solvents.
- Evaluating dose- and time-dependent cytotoxicity induced by emodin.
Main Results:
- Emodin's absorption spectrum is solvent-dependent, with water-containing solvents causing a red shift and increased overlap with formazan absorption.
- Emodin forms formazan in a dose-dependent manner, partially suppressed by serum.
- Emodin induces cytotoxicity in a time- and dose-dependent manner.
- Emodin alters the accuracy of the standard MTT assay.
Conclusions:
- Emodin can interfere with the accuracy of the MTT assay.
- A modified MTT assay remains a valuable tool for quantifying emodin-induced cytotoxicity despite potential interferences.
