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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Main Authors: Ratih Asmana Ningrum, Yulaika Romadhani, Popi Hadi Wisnuwardhani, Adi Santoso
Format: Article
Language:English
Published: Universitas Gadjah Mada 2014-10-01
Series:Indonesian Journal of Pharmacy
Subjects:
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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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
work_keys_str_mv AT ratihasmananingrum optimizationofsodiumdodecylsulphateasaformazansolventandcomparisonof345dimethylthiazo2yl25diphenyltetrazoliumbromidemttassaywithwst1assayinmcf7cells
AT yulaikaromadhani optimizationofsodiumdodecylsulphateasaformazansolventandcomparisonof345dimethylthiazo2yl25diphenyltetrazoliumbromidemttassaywithwst1assayinmcf7cells
AT popihadiwisnuwardhani optimizationofsodiumdodecylsulphateasaformazansolventandcomparisonof345dimethylthiazo2yl25diphenyltetrazoliumbromidemttassaywithwst1assayinmcf7cells
AT adisantoso optimizationofsodiumdodecylsulphateasaformazansolventandcomparisonof345dimethylthiazo2yl25diphenyltetrazoliumbromidemttassaywithwst1assayinmcf7cells