OPTIMIZATION OF SODIUM DODECYL SULPHATE AS A FORMAZAN SOLVENT AND COMPARISON OF 3-(4,-5- DIMETHYLTHIAZO-2-YL)-2,5-DIPHENYLTETRAZOLIUM BROMIDE (MTT) ASSAY WITH WST-1 ASSAY IN MCF-7 CELLS
A 3-(4,-5-dimethylthiazo-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay is a method that used to measure cell viability. It is based on the conversion of MTT by succinic dehydrogenase enzyme into insoluble formazan. Dissolution of formazan by using proper solvent is the most important step of...
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Universitas Gadjah Mada
2014-10-01
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Series: | Indonesian Journal of Pharmacy |
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Online Access: | http://indonesianjpharm.farmasi.ugm.ac.id/index.php/3/article/view/274 |
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author | Ratih Asmana Ningrum Yulaika Romadhani Popi Hadi Wisnuwardhani Adi Santoso |
author_facet | Ratih Asmana Ningrum Yulaika Romadhani Popi Hadi Wisnuwardhani Adi Santoso |
author_sort | Ratih Asmana Ningrum |
collection | DOAJ |
description | A 3-(4,-5-dimethylthiazo-2-yl)-2,5-diphenyltetrazolium bromide
(MTT) assay is a method that used to measure cell viability. It is
based on the conversion of MTT by succinic dehydrogenase enzyme
into insoluble formazan. Dissolution of formazan by using proper
solvent is the most important step of MTT assay to obtain valid and
reliable data. In this study, we observed several solvents
[isopropanol, dimethyl sulfoxide (DMSO), and sodium dodecyl
sulphate (SDS)] to validate MTT assay by using MCF-7 cells. The
observation was performed by MTT addition at concentration of
0.5µg/µL, 3-4h cells incubation at 37°C, dissolution of formed
formazan crystal and absorbance measurement at 570nm. The
result showed that formazan completely dissolved in DMSO and
10% SDS. The most advantage of using SDS was it avoided the
removal of partially dissolved formazan. In this observation, we
also found that pH was a very important factor in SDS solution that
affected the reaction. The use of optimal condition on MTT assay by
SDS-0.01M HCl and SDS-0.025M HCl as formazan solvents showed
that IC50 of curcumin were 32.3±0.78µM and 24.08±1.72µM
respectively, while WST-1 assay resulted IC50 of curcumin
80.69±5.35µM. Altogether, this study strongly indicated that SDS0.01M
HCl was the best formazan solvent for MTT assay. |
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id | doaj.art-9e9b0a0789a841a3a34ff63182f98f72 |
institution | Directory Open Access Journal |
issn | 2338-9427 2338-9486 |
language | English |
last_indexed | 2024-12-20T21:27:26Z |
publishDate | 2014-10-01 |
publisher | Universitas Gadjah Mada |
record_format | Article |
series | Indonesian Journal of Pharmacy |
spelling | doaj.art-9e9b0a0789a841a3a34ff63182f98f722022-12-21T19:26:08ZengUniversitas Gadjah MadaIndonesian Journal of Pharmacy2338-94272338-94862014-10-0125424525410.14499/indonesianjpharm25iss4pp245OPTIMIZATION OF SODIUM DODECYL SULPHATE AS A FORMAZAN SOLVENT AND COMPARISON OF 3-(4,-5- DIMETHYLTHIAZO-2-YL)-2,5-DIPHENYLTETRAZOLIUM BROMIDE (MTT) ASSAY WITH WST-1 ASSAY IN MCF-7 CELLS Ratih Asmana Ningrum0Yulaika Romadhani1Popi Hadi Wisnuwardhani2Adi Santoso3Research Center for Biotechnology, Indonesian Institute of Sciences Jl. Raya Bogor KM 46, Cibinong, Bogor 16911 IndonesiaResearch Center for Biotechnology, Indonesian Institute of Sciences Jl. Raya Bogor KM 46, Cibinong, Bogor 16911 IndonesiaResearch Center for Biotechnology, Indonesian Institute of Sciences Jl. Raya Bogor KM 46, Cibinong, Bogor 16911 IndonesiaResearch Center for Biotechnology, Indonesian Institute of Sciences Jl. Raya Bogor KM 46, Cibinong, Bogor 16911 IndonesiaA 3-(4,-5-dimethylthiazo-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay is a method that used to measure cell viability. It is based on the conversion of MTT by succinic dehydrogenase enzyme into insoluble formazan. Dissolution of formazan by using proper solvent is the most important step of MTT assay to obtain valid and reliable data. In this study, we observed several solvents [isopropanol, dimethyl sulfoxide (DMSO), and sodium dodecyl sulphate (SDS)] to validate MTT assay by using MCF-7 cells. The observation was performed by MTT addition at concentration of 0.5µg/µL, 3-4h cells incubation at 37°C, dissolution of formed formazan crystal and absorbance measurement at 570nm. The result showed that formazan completely dissolved in DMSO and 10% SDS. The most advantage of using SDS was it avoided the removal of partially dissolved formazan. In this observation, we also found that pH was a very important factor in SDS solution that affected the reaction. The use of optimal condition on MTT assay by SDS-0.01M HCl and SDS-0.025M HCl as formazan solvents showed that IC50 of curcumin were 32.3±0.78µM and 24.08±1.72µM respectively, while WST-1 assay resulted IC50 of curcumin 80.69±5.35µM. Altogether, this study strongly indicated that SDS0.01M HCl was the best formazan solvent for MTT assay.http://indonesianjpharm.farmasi.ugm.ac.id/index.php/3/article/view/274DMSOMCF-7 cellsMTT assaysodium dodecyl sulphateWST-1 assay |
spellingShingle | Ratih Asmana Ningrum Yulaika Romadhani Popi Hadi Wisnuwardhani Adi Santoso OPTIMIZATION OF SODIUM DODECYL SULPHATE AS A FORMAZAN SOLVENT AND COMPARISON OF 3-(4,-5- DIMETHYLTHIAZO-2-YL)-2,5-DIPHENYLTETRAZOLIUM BROMIDE (MTT) ASSAY WITH WST-1 ASSAY IN MCF-7 CELLS Indonesian Journal of Pharmacy DMSO MCF-7 cells MTT assay sodium dodecyl sulphate WST-1 assay |
title | OPTIMIZATION OF SODIUM DODECYL SULPHATE AS A FORMAZAN SOLVENT AND COMPARISON OF 3-(4,-5- DIMETHYLTHIAZO-2-YL)-2,5-DIPHENYLTETRAZOLIUM BROMIDE (MTT) ASSAY WITH WST-1 ASSAY IN MCF-7 CELLS |
title_full | OPTIMIZATION OF SODIUM DODECYL SULPHATE AS A FORMAZAN SOLVENT AND COMPARISON OF 3-(4,-5- DIMETHYLTHIAZO-2-YL)-2,5-DIPHENYLTETRAZOLIUM BROMIDE (MTT) ASSAY WITH WST-1 ASSAY IN MCF-7 CELLS |
title_fullStr | OPTIMIZATION OF SODIUM DODECYL SULPHATE AS A FORMAZAN SOLVENT AND COMPARISON OF 3-(4,-5- DIMETHYLTHIAZO-2-YL)-2,5-DIPHENYLTETRAZOLIUM BROMIDE (MTT) ASSAY WITH WST-1 ASSAY IN MCF-7 CELLS |
title_full_unstemmed | OPTIMIZATION OF SODIUM DODECYL SULPHATE AS A FORMAZAN SOLVENT AND COMPARISON OF 3-(4,-5- DIMETHYLTHIAZO-2-YL)-2,5-DIPHENYLTETRAZOLIUM BROMIDE (MTT) ASSAY WITH WST-1 ASSAY IN MCF-7 CELLS |
title_short | OPTIMIZATION OF SODIUM DODECYL SULPHATE AS A FORMAZAN SOLVENT AND COMPARISON OF 3-(4,-5- DIMETHYLTHIAZO-2-YL)-2,5-DIPHENYLTETRAZOLIUM BROMIDE (MTT) ASSAY WITH WST-1 ASSAY IN MCF-7 CELLS |
title_sort | optimization of sodium dodecyl sulphate as a formazan solvent and comparison of 3 4 5 dimethylthiazo 2 yl 2 5 diphenyltetrazolium bromide mtt assay with wst 1 assay in mcf 7 cells |
topic | DMSO MCF-7 cells MTT assay sodium dodecyl sulphate WST-1 assay |
url | http://indonesianjpharm.farmasi.ugm.ac.id/index.php/3/article/view/274 |
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