Sensitivity of two-stage PCR amplification for detection of Mycobacterium tuberculosis in paraffin-embedded tissues

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dc.authoridGünal, Selami/0000-0002-4752-5176
dc.authoridDURMAZ, RIZA/0000-0001-6561-778X
dc.authoridAYDIN, Nasuhi/0000-0003-3145-2432
dc.authorwosidAydin, Nasuhi E/B-6536-2012
dc.authorwosidAYDIN, Nasuhi Engin/L-1607-2019
dc.authorwosidGünal, Selami/AAA-7246-2021
dc.authorwosidAYDIN, Nasuhi/O-9043-2016
dc.contributor.authorDurmaz, R
dc.contributor.authorAydin, A
dc.contributor.authorDurmaz, B
dc.contributor.authorAydin, NE
dc.contributor.authorAkbasak, BS
dc.contributor.authorGunal, S
dc.date.accessioned2024-08-04T20:11:52Z
dc.date.available2024-08-04T20:11:52Z
dc.date.issued1997
dc.departmentİnönü Üniversitesien_US
dc.description.abstractIn order to improve the sensitivity of polymerase chain reaction (PCR) for the detection of mycobacterial DNA in paraffin-embedded tissues, a new approach with two sets of specific primers in two-stage PCR was employed in specimens obtained from tuberculosis patients and controls. Thirty-nine paraffin blocks selected from patients who had been diagnosed as having tuberculosis by radiological evaluations, histopathological findings,land clinical symptoms and signs including response to antituberculous treatment were examined. The control group consisted of 10 specimens from individuals that were proved to be negative for tuberculosis. After deparaffinization, lysis, phenol-chloroform extraction, and ethanol precipitation, the isolated DNA was amplified by PCR. Initially, all specimens were examined by the one-stage PCR using specific primers for 123-base pair (bp) fragment in IS6110 of mycobacterial DNA which yielded positive results only in 3 out of 39 (7.7%). In the two-stage PCR technique, 245-bp fragment of mycobacterial DNA was amplified at the first-step, then the PCR products were reamplified using the second specific primer pairs for 123-bp fragment. The true positivity of the two-stage PCR was 84.6% (33/39). The results indicate that two-stage PCR is more sensitive than one-stage (84.6% vs. 7.7%). All control specimens were negative by both PCR amplification methods, indicating that specificity of both methods was high. When the two-stage amplification was used, PCR positivity in the specimens obtained from different tissues was as follows: peritoneal and omental biopsies, 4/4; bone biopsies, 3/3; lymph node biopsies, 12/14; genito-urinary biopsies, 7/9; skin biopsies, 4/6; and one from each lung, breast, and pleural biopsies. PCR showed a good correlation with the granulomatous tissue reaction resulting in a 83.8% (31/37) positivity. The results indicate that the two-stage PCR amplification can be used for detection of M. tuberculosis in paraffin-embedded tissues and is a useful technique in confirming tuberculosis in patients with clinically suspected disease who have acid-fast stain-negative. (C) 1997 Elsevier Science B.V.en_US
dc.identifier.doi10.1016/S0167-7012(97)00022-5
dc.identifier.endpage75en_US
dc.identifier.issn0167-7012
dc.identifier.issue2en_US
dc.identifier.scopus2-s2.0-0030837768en_US
dc.identifier.scopusqualityN/Aen_US
dc.identifier.startpage69en_US
dc.identifier.urihttps://doi.org/10.1016/S0167-7012(97)00022-5
dc.identifier.urihttps://hdl.handle.net/11616/93063
dc.identifier.volume29en_US
dc.identifier.wosWOS:A1997XM16300001en_US
dc.identifier.wosqualityQ3en_US
dc.indekslendigikaynakWeb of Scienceen_US
dc.indekslendigikaynakScopusen_US
dc.language.isoenen_US
dc.publisherElsevier Science Bven_US
dc.relation.ispartofJournal of Microbiological Methodsen_US
dc.relation.publicationcategoryMakale - Uluslararası Hakemli Dergi - Kurum Öğretim Elemanıen_US
dc.rightsinfo:eu-repo/semantics/closedAccessen_US
dc.subjectM-tuberculosisen_US
dc.subjectparaffin blocken_US
dc.subjecttwo-stage PCRen_US
dc.titleSensitivity of two-stage PCR amplification for detection of Mycobacterium tuberculosis in paraffin-embedded tissuesen_US
dc.typeArticleen_US

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