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2012-03-13Zeitschriftenartikel DOI: 10.3109/13693786.2012.665613
Enhanced fungal DNA-extraction from formalin-fixed, paraffin-embedded tissue specimens by application of thermal energy
dc.contributor.authorRickerts, Volker
dc.contributor.authorKhot, Prasanna D.
dc.contributor.authorKo, Daisy L.
dc.contributor.authorFredricks, David N.
dc.date.accessioned2018-05-07T16:17:27Z
dc.date.available2018-05-07T16:17:27Z
dc.date.created2013-03-25
dc.date.issued2012-03-13none
dc.identifier.otherhttp://edoc.rki.de/oa/articles/reMKZJ8wZmDQ/PDF/23Wo7ZEApET9w.pdf
dc.identifier.urihttp://edoc.rki.de/176904/1425
dc.description.abstractDetermining the etiology of invasive fungal infections (IFI) is critical for patient management as fungi vary in their susceptibility to antifungals. However, the etiology remains obscure in many cases due to negative culture results. The identification of fungal DNA by PCR in pathology blocks and sequencing it is an alternative approach to determine the cause of IFI. Previous studies identified fungal DNA in only 50% of samples with positive histopathology results, probably due to DNA damage by tissue fixation. We used realtime PCR to quantify human and fungal DNA from formalin-fixed, paraffin-embedded tissue specimens in order to study the effect of thermal energy during extraction on the yield of amplifiable DNA and subsequent identification of fungal DNA. Tissue sections from eight patients with proven IFI were subjected to DNA extraction with varying exposure to thermal energy. Amplifiable DNA increased up to 76-fold by increasing the incubation temperature from 65°C to 90°C and an additional increase was documented by incubating samples for up to 6 hours at this temperature. The augmented amplification of fungal DNA was associated with improved species identification by the sequencing of the PCR amplicons. This may help illuminate the etiology of IFI and thereby improve patient management by guiding antifungal therapy.eng
dc.language.isoeng
dc.publisherRobert Koch-Institut, Infektionskrankheiten / Erreger
dc.subjectHumanseng
dc.subjectSensitivity and Specificityeng
dc.subjectPolymerase Chain Reaction/methodseng
dc.subjectHot Temperatureeng
dc.subjectAspergillosis/microbiologyeng
dc.subjectAspergillus/geneticseng
dc.subjectAspergillus/isolation & purificationeng
dc.subjectCandida/geneticseng
dc.subjectCandida/isolation & purificationeng
dc.subjectDNA Fungal/isolation & purificationeng
dc.subjectFormaldehyde/chemistryeng
dc.subjectGenome Humaneng
dc.subjectLung/microbiologyeng
dc.subjectNucleic Acid Amplificationeng
dc.subjectTechniques/methodseng
dc.subjectParaffin Embeddingeng
dc.subjectPathology Molecular/methodseng
dc.subjectTissue Fixation/methodseng
dc.subject.ddc610 Medizin
dc.titleEnhanced fungal DNA-extraction from formalin-fixed, paraffin-embedded tissue specimens by application of thermal energy
dc.typeperiodicalPart
dc.identifier.urnurn:nbn:de:0257-10029694
dc.identifier.doi10.3109/13693786.2012.665613
dc.identifier.doihttp://dx.doi.org/10.25646/1350
local.edoc.container-titleMedical Mycology
local.edoc.fp-subtypeArtikel
local.edoc.type-nameZeitschriftenartikel
local.edoc.container-typeperiodical
local.edoc.container-type-nameZeitschrift
local.edoc.container-urlhttp://informahealthcare.com/doi/abs/10.3109/13693786.2012.665613
local.edoc.container-publisher-nameInforma Healthcare
local.edoc.container-volume50
local.edoc.container-issue6
local.edoc.container-year2012

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