Parallel detection of five human herpes virus DNAs by a set of real-time polymerase chain reactions in a single run

Stöcher, Markus; Leb, Victoria; Bozic, Michael; Kessler, Harald H.; Halwachs‐Baumann, Gabriele; Landt, Olfert; Stekel, Herbert; Berg, Jörg

doi:10.1016/s1386-6532(02)00042-2

Cited by 58 publications

(45 citation statements)

References 19 publications

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“…Using real-time PCR, most of the problems associated with PCR (contamination, cumbersome detection, and rather expensive tests) are solved, and a rapid, economical (considering the reduced workload), andmost importantly -closed system is available. The use of internal controls simplified the assay protocol and allowed monitoring of sample adequacy (10,22,23).…”

Section: Discussionmentioning

confidence: 99%

Diagnosis of infection with human herpes viruses in routine laboratory practice

Mostafaie

Huber²,

Sebesta³

et al. 2009

Clinical Chemistry and Laboratory Medicine

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Background: The potential for faster detection of human herpes viruses using PCR compared to other methods is undisputed. However, because of fear of contamination, the clinical implication of nucleic amplification methods in routine laboratories is not widespread. Herpes viruses cause a wide spectrum of diseases and can cause morbidity and mortality in immune-compromised patients. Using real-time PCR, most of the problems associated with PCR (contamination, cumbersome detection, and rather expensive tests) are solved, and a rapid, economical, and -most importantly -closed system is at hand. Methods: We evaluated work procedures in our laboratory that enable the routine diagnosis of viral infections with high accuracy and rapid turn-around time. In parallel, inherent problems usually associated with PCR testing, especially cross-contamination could be suppressed to a minimum. The start of the work flow process begins with an automated nucleic acid extraction procedure that yields high quality DNA. A common -internally and externally controlled -PCR program for all six viruses allows rapid sample turn around. Results: In all, 7500 analyses for human herpes virus infection were performed in the last 5 years. Results for various different specimens were produced within 24 h. Contamination occurred rarely and could be ameliorated easily. The use of internal controls identified rare PCR-inhibited samples. The detection limits

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Section: Discussionmentioning

confidence: 99%

Diagnosis of infection with human herpes viruses in routine laboratory practice

Mostafaie

Huber²,

Sebesta³

et al. 2009

Clinical Chemistry and Laboratory Medicine

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“…Most of these assays use specific targets, such as UL83, UL123 genes or the HXFL4 region (Alain et al;Caliendo et al, 2007;Gault et al, 2001;Gouarin et al, 2007;Mengelle et al, 2003). The most common targets used for the detection of CMV by RT-PCR are the immediate early (IE) gene (Nitsche et al, 1999), the polymerase (UL54) gene (Schaade et al, 2000) , the glycoprotein B gene (UL55) (Espy et al, 2006) and the pp65 gene (UL83) (Stocher et al, 2003). Among the commercially available standardized methods to detect CMV infection, the LightCycler® CMV Quantitative Kit (Roche), the RealArt CMV LightCycler PCR reagent test (QIAGEN, Germantown, MD), CMV R-gene TM (Argene, France), Affigene CMV Trender (Cepheid, Sweden) and the Abbott CMV real-time PCR Kit (Abbott Diagnosis, USA) have been evaluated in SCT.…”

Section: Commercial Rt-pcr Assaysmentioning

confidence: 99%

Detection of CMV Infection in Allogeneic SCT Recipients: The Multiple Assays

Lobo¹,

BenMarzouk-Hidalgo²,

Pérez‐Romero³

2012

Advances in Hematopoietic Stem Cell Research

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“…Virus specific DNA sequences for HSV were adapted from the genes quantitatively amplified by Stocher et al 13 The probe length was extended by flanking sequences of the virus genome to obtain a 37 basepair target sequence with complementary capture and reporter probes ( Table 2). The sequences were analyzed with IDT SciTools Oligoanalyzer 14 to determine the secondary structure of hairpins and homodimers.…”

Section: Probe and Target Sequencesmentioning

confidence: 99%

Amplification free detection of Herpes Simplex Virus DNA

Thomson¹,

Dimitrov²,

Cooper³

2011

Analyst

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Amplification-free detection of nucleic acids in complex biological samples is an important technology for clinical diagnostics, especially in the case where the detection is quantitative and highly sensitive. Here we present the detection of a synthetic DNA sequence from Herpes Simplex Virus-1 within swine cerebrospinal fluid (CSF), using a sandwich-like, magnetic nanoparticle pull-down assay. Magnetic nanoparticles and fluorescent polystyrene nanoparticles were both modified with DNA probes, able to hybridise either end of the target DNA, forming the sandwich-like complex which can be captured magnetically and detected by fluorescence. The concentration of the target DNA was determined by counting individual and aggregated fluorescent nanoparticles on a planar glass surface within a fluidic chamber. DNA probe coupling for both nanoparticles was optimized. Polystyrene reporter nanoparticles that had been modified with amine terminated DNA probes were also treated with amine terminated polyethylene glycol, in order to reduce non-specific aggregation and target independent adhesion to the magnetic particles. This way, a limit of detection for the target DNA of 0.8 pM and 1 pM could be achieved for hybridisation buffer and CSF respectively, corresponding to 0.072 and 0.090 femtomoles of target DNA, in a volume of 0.090 mL.

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Parallel detection of five human herpes virus DNAs by a set of real-time polymerase chain reactions in a single run

Cited by 58 publications

References 19 publications

Diagnosis of infection with human herpes viruses in routine laboratory practice

Diagnosis of infection with human herpes viruses in routine laboratory practice

Detection of CMV Infection in Allogeneic SCT Recipients: The Multiple Assays

Amplification free detection of Herpes Simplex Virus DNA

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