ArticleReports of biochemistry & molecular biology2013
Quantification and optimization of Candida albicans DNA in blood samples using Real- Time PCR.
Article in Reports of biochemistry & molecular biology, 2013. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 4 papers.
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Who cites it
4 citing papers in PubMed, 7 citations in OpenAlex.
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- Fluorescent toys 'n' tools lighting the way in fungal research.FEMS microbiology reviews · 2021Article
- PCR-detectableOpen life sciences · 2020Article
- A real time PCR assay on blood for diagnosis of invasive candidiasis in immunocompromised patient.Current medical mycology · 2015Article
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Authors and funding
6 authors at 2 institutions in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
backgroundCandida albicans (C. albicans) is a major cause of candidaemia in people with impaired immunity. Blood culture is a "gold standard" for candidaemia detection but is time-consuming and relatively insensitive. We established a real-time PCR assay for C. albicans detection in blood by LightCycler PCR and melting curve analysis.
methodsFive milliliter blood samples from healthy volunteers were spiked with 10(0)-10(6) C. albicans cells to determine the detection limit of our method. DNA was extracted from whole blood using glass beads and the QIAamp DNA Blood Mini Kit (Qiagen, Hilden Germany). DNA from C. albicans isolates were amplified with primers and inserted into Escherichia coli (E. coli) DH5α.1 cells with the TA cloning vector (Invitrogen). The plasmid was used for standardization and optimization. A quantitative PCR assay with the LightCycler amplification and detection system based on fluorescence resonance energy transfer (FRET) with two different specific probes was established. To assess the precision and reproducibility of real-time PCR the intra-assay precision was determined in six consecutive assays.
resultsNo cross-reactivity of the hybridization probes with the DNA of non-C. albicans species or human genomic DNA was observed, which confirmed its 100% specificity. The minimum limit detected was one C. albicans cell or 10(0) CFU/ml (10 fg) per PCR reaction. The real-time PCR efficiency rate for Candida was high (E = 1.95). Melting curve analysis of C. albicans showed a specific melting peak temperature of 65.76 °C.
conclusionThe real-time PCR assay we developed is highly specific and sufficiently sensitive to detect the fungal load for early diagnosis of invasive candidiasis.
Indexed as
Identifiers
26989719PMC4757066W2276253049What OpenQuestion holds
Registered trials
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