Trypan blue exclusion assay by flow cytometry
| Autor(a) principal: | |
|---|---|
| Data de Publicação: | 2014 |
| Outros Autores: | , , , , , , |
| Tipo de documento: | Artigo |
| Idioma: | eng |
| Título da fonte: | Repositório Institucional da FIOCRUZ (ARCA) |
| Texto Completo: | https://www.arca.fiocruz.br/handle/icict/9397 |
Resumo: | Universidade Federal dos Vales do Jequitinhonha e Mucuri. Departamento de Farmácia. Laboratório de Imunologia. Diamantina, MG, Brasil. |
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Freitas, Bethânia Alves de AvelarAlmeida, Valéria GomesPinto, Mauro Cunha XavierMourão, Flávio Afonso GonçalvesMassensini, Andre RicardoMartins Filho, Olindo AssisVieira, Etel RochaMelo, Gustavo Eustáquio Brito Alvim de2015-01-28T15:33:09Z2015-01-28T15:33:09Z2014FREITAS, Bethânia Alves de Avelar, et al. Trypan blue exclusion assay by flow cytometry. Braz J Med Biol Res; v. 47, n. 4, p. 307-15, 20140100-879Xhttps://www.arca.fiocruz.br/handle/icict/939710.1590/1414-431X20143437engAssociação Brasileira de Divulgação CientíficaTrypan blue exclusion assay by flow cytometryinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articleUniversidade Federal dos Vales do Jequitinhonha e Mucuri. Departamento de Farmácia. Laboratório de Imunologia. Diamantina, MG, Brasil.Universidade Federal dos Vales do Jequitinhonha e Mucuri. Departamento de Farmácia. Laboratório de Imunologia. Diamantina, MG, BrasilUniversidade Federal de Minas Gerais. Instituto de Ciências Biológicas. Departamento de Fisiologia e Farmacologia. Belo Horizonte, MG, Brasil.Universidade Federal de Minas Gerais. Instituto de Ciências Biológicas. Departamento de Fisiologia e Farmacologia. Belo Horizonte, MG, Brasil.Universidade Federal de Minas Gerais. Instituto de Ciências Biológicas. Departamento de Fisiologia e Farmacologia. Belo Horizonte, MG, Brasil.Fundação Osvaldo Cruz. Centro de Pesquisas René Rachou. Laboratório de Biomarcadores de Diagnóstico e Monitoração Belo Horizonte, MG, Brasil.Universidade Federal dos Vales do Jequitinhonha e Mucuri. Departamento de Farmácia. Laboratório de Imunologia. Diamantina, MG, Brasil.Universidade Federal dos Vales do Jequitinhonha e Mucuri. Departamento de Farmácia. Laboratório de Imunologia. Diamantina, MG, Brasil.Dye exclusion tests are used to determine the number of live and dead cells. These assays are based on the principle that intact plasma membranes in live cells exclude specific dyes, whereas dead cells do not. Although widely used, the trypan blue (TB) exclusion assay has limitations. The dye can be incorporated by live cells after a short exposure time, and personal reliability, related to the expertise of the analyst, can affect the results. We propose an alternative assay for evaluating cell viability that combines the TB exclusion test and the high sensitivity of the flow cytometry technique. Previous studies have demonstrated the ability of TB to emit fluorescence when complexed with proteins. According to our results, TB/bovine serum albumin and TB/cytoplasmic protein complexes emit fluorescence at 660 nm, which is detectable by flow cytometry using a 650-nm low-pass band filter. TB at 0.002% (w/v) was defined as the optimum concentration for distinguishing unstained living cells from fluorescent dead cells, and fluorescence emission was stable for 30 min after cell treatment. Although previous studies have shown that TB promotes green fluorescence quenching, TB at 0.002% did not interfere with green fluorescence in human live T-cells stained with anti-CD3/fluorescein isothiocyanate (FITC) monoclonal antibody. We observed a high correlation between the percentage of propidium iodide+CD3/FITC+ and TB+CD3/FITC+ cells, as well as similar double-stained cell profiles in flow cytometry dot-plot graphs. Taken together, the results indicate that a TB exclusion assay by flow cytometry can be employed as an alternative tool for quick and reliable cell viability analysis.Trypan blueFluorescenceFlow cytometryCytotoxicityinfo:eu-repo/semantics/openAccessreponame:Repositório Institucional da FIOCRUZ (ARCA)instname:Fundação Oswaldo Cruz (FIOCRUZ)instacron:FIOCRUZLICENSElicense.txtlicense.txttext/plain; charset=utf-81914https://www.arca.fiocruz.br/bitstream/icict/9397/1/license.txt7d48279ffeed55da8dfe2f8e81f3b81fMD51ORIGINAL2014_003.pdf2014_003.pdfapplication/pdf1135145https://www.arca.fiocruz.br/bitstream/icict/9397/2/2014_003.pdfdc2e947d746266f16712e0890b86299cMD52TEXT2014_003.pdf.txt2014_003.pdf.txtExtracted texttext/plain36197https://www.arca.fiocruz.br/bitstream/icict/9397/3/2014_003.pdf.txtafa7fd3d60b3df49b91addb07b91f3b0MD53icict/93972019-06-19 10:07:35.846oai:www.arca.fiocruz.br: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ório InstitucionalPUBhttps://www.arca.fiocruz.br/oai/requestrepositorio.arca@fiocruz.bropendoar:21352019-06-19T13:07:35Repositório Institucional da FIOCRUZ (ARCA) - Fundação Oswaldo Cruz (FIOCRUZ)false |
| dc.title.pt_BR.fl_str_mv |
Trypan blue exclusion assay by flow cytometry |
| title |
Trypan blue exclusion assay by flow cytometry |
| spellingShingle |
Trypan blue exclusion assay by flow cytometry Freitas, Bethânia Alves de Avelar Trypan blue Fluorescence Flow cytometry Cytotoxicity |
| title_short |
Trypan blue exclusion assay by flow cytometry |
| title_full |
Trypan blue exclusion assay by flow cytometry |
| title_fullStr |
Trypan blue exclusion assay by flow cytometry |
| title_full_unstemmed |
Trypan blue exclusion assay by flow cytometry |
| title_sort |
Trypan blue exclusion assay by flow cytometry |
| author |
Freitas, Bethânia Alves de Avelar |
| author_facet |
Freitas, Bethânia Alves de Avelar Almeida, Valéria Gomes Pinto, Mauro Cunha Xavier Mourão, Flávio Afonso Gonçalves Massensini, Andre Ricardo Martins Filho, Olindo Assis Vieira, Etel Rocha Melo, Gustavo Eustáquio Brito Alvim de |
| author_role |
author |
| author2 |
Almeida, Valéria Gomes Pinto, Mauro Cunha Xavier Mourão, Flávio Afonso Gonçalves Massensini, Andre Ricardo Martins Filho, Olindo Assis Vieira, Etel Rocha Melo, Gustavo Eustáquio Brito Alvim de |
| author2_role |
author author author author author author author |
| dc.contributor.author.fl_str_mv |
Freitas, Bethânia Alves de Avelar Almeida, Valéria Gomes Pinto, Mauro Cunha Xavier Mourão, Flávio Afonso Gonçalves Massensini, Andre Ricardo Martins Filho, Olindo Assis Vieira, Etel Rocha Melo, Gustavo Eustáquio Brito Alvim de |
| dc.subject.en.pt_BR.fl_str_mv |
Trypan blue Fluorescence Flow cytometry Cytotoxicity |
| topic |
Trypan blue Fluorescence Flow cytometry Cytotoxicity |
| description |
Universidade Federal dos Vales do Jequitinhonha e Mucuri. Departamento de Farmácia. Laboratório de Imunologia. Diamantina, MG, Brasil. |
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2014 |
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2014 |
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2015-01-28T15:33:09Z |
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2015-01-28T15:33:09Z |
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info:eu-repo/semantics/publishedVersion |
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info:eu-repo/semantics/article |
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FREITAS, Bethânia Alves de Avelar, et al. Trypan blue exclusion assay by flow cytometry. Braz J Med Biol Res; v. 47, n. 4, p. 307-15, 2014 |
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0100-879X |
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10.1590/1414-431X20143437 |
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FREITAS, Bethânia Alves de Avelar, et al. Trypan blue exclusion assay by flow cytometry. Braz J Med Biol Res; v. 47, n. 4, p. 307-15, 2014 0100-879X 10.1590/1414-431X20143437 |
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