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Colorimetric microwell plate reverse-hybridization assay for Mycobacterium tuberculosis detection

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Colorimetric microwell plate reverse-hybridization assay for Mycobacterium tuberculosis detection

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Título Colorimetric microwell plate reverse-hybridization assay for Mycobacterium tuberculosis detection
Autor Michelon, Candice Tosi
Rosso, Franciele
Schmid, Karen Barros
Sperhacke, Rosa Dea
Oliveira, Martha Maria
Kritski, Afrânio Lineu
Rezende Junior, Leonides
Costa, Elis Regina Dalla
Ribeiro, Andrezza Wolowski
Verza, Mirela
Cafrune, Patricia Izquierdo
Silva, Marcia Susana Nunes
Kühleis, Daniele Chaves
Zaha, Arnaldo
Rossetti, Maria Lucia Rosa
Abstract Direct smear examination using Ziehl-Neelsen staining for pulmonary tuberculosis (PTB) diagnosis is inexpensive and easy to use, but has the major limitation of low sensitivity. Rapid molecular methods are becoming more widely available in centralized laboratories, but they depend on timely reporting of results and strict quality assurance obtainable only from costly commercial kits available in high burden nations. This study describes a pre-commercial colorimetric method, Detect-TB, for detecting Mycobacterium tuberculosis DNA in which an oligonucleotide probe is fixed onto wells of microwell plates and hybridized with biotinylated polymerase chain reaction amplification products derived from clinical samples. The probe is capable of hybridising with the IS6110 insertion element and was used to specifically recognise the M. tuberculosis complex. When combined with an improved silica-based DNA extraction method, the sensitivity of the test was 50 colony-forming units of the M. tuberculosis reference strain H37Rv. The results that were in agreement with reference detection methods were observed in 95.2% (453/476) of samples included in the analysis. Sensitivity and specificity for 301 induced sputum samples and 175 spontaneous sputum samples were 85% and 98%, and 94% and 100%, respectively. This colorimetric method showed similar specificity to that described for commercially available kits and may provide an important contribution for PTB diagnosis.
Contido em Memorias do Instituto Oswaldo Cruz : an International Journal of Biological and Biomedical Research. Rio de Janeiro. Vol. 106, n. 2 (mar. 2011), p. 194-199
Assunto Hibridização
Mycobacterium tuberculosis
Tuberculose
[en] Hybridization assay
[en] IS6110
[en] Molecular diagnosis
[en] Oligonucleotide probe
[en] Reverse
[en] Tuberculosis
Origem Nacional
Tipo Artigo de periódico
URI http://hdl.handle.net/10183/30533
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