Calibrated real-time PCR assay for quantitation of human herpesvirus 8 DNA in biological fluids

Francesco Broccolo, Giuseppe Locatelli, Loredana Sarmati, Sara Piergiovanni, Fabrizio Veglia, Massimo Andreoni, Stefano Buttò, Barbara Ensoli, Paolo Lusso, Mauro S. Malnati

Research output: Contribution to journalArticle

40 Citations (Scopus)

Abstract

Accurate laboratory tests for the diagnosis of active human herpesvirus 8 (HHV-8) infection are becoming essential to study the pathogenesis of HHV-8-associated tumors and for the clinical management of HHV-8-infected individuals. We have developed a highly sensitive, calibrated quantitative real-time PCR assay for the measurement of cell-free HHV-8 DNA in body fluids, based on the addition of a synthetic DNA calibrator prior to DNA extraction. The calibrator controls each sample for the presence of PCR inhibitors, determines a cutoff value of sensitivity for negative samples, and normalizes positive samples for the efficiency of DNA recovery. The assay shows a wide dynamic range of detection (between 1 and 106 viral genome equivalents/reaction) and a high degree of accuracy even in the presence of high amounts (up to 1 μg) of human genomic DNA. Moreover, the assay has a very high sensitivity (lower detection limit, 10 genome equivalents/ml) and a high degree of reproducibility and repeatability with a coefficient of variation (CV) of

Original languageEnglish
Pages (from-to)4652-4658
Number of pages7
JournalJournal of Clinical Microbiology
Volume40
Issue number12
DOIs
Publication statusPublished - Dec 1 2002

Fingerprint

Human Herpesvirus 8
Real-Time Polymerase Chain Reaction
DNA
Herpesviridae Infections
Clinical Laboratory Techniques
Viral Genome
Body Fluids
Limit of Detection
Genome
Polymerase Chain Reaction
Neoplasms

ASJC Scopus subject areas

  • Microbiology (medical)
  • Microbiology

Cite this

Calibrated real-time PCR assay for quantitation of human herpesvirus 8 DNA in biological fluids. / Broccolo, Francesco; Locatelli, Giuseppe; Sarmati, Loredana; Piergiovanni, Sara; Veglia, Fabrizio; Andreoni, Massimo; Buttò, Stefano; Ensoli, Barbara; Lusso, Paolo; Malnati, Mauro S.

In: Journal of Clinical Microbiology, Vol. 40, No. 12, 01.12.2002, p. 4652-4658.

Research output: Contribution to journalArticle

Broccolo, F, Locatelli, G, Sarmati, L, Piergiovanni, S, Veglia, F, Andreoni, M, Buttò, S, Ensoli, B, Lusso, P & Malnati, MS 2002, 'Calibrated real-time PCR assay for quantitation of human herpesvirus 8 DNA in biological fluids', Journal of Clinical Microbiology, vol. 40, no. 12, pp. 4652-4658. https://doi.org/10.1128/JCM.40.12.4652-4658.2002
Broccolo, Francesco ; Locatelli, Giuseppe ; Sarmati, Loredana ; Piergiovanni, Sara ; Veglia, Fabrizio ; Andreoni, Massimo ; Buttò, Stefano ; Ensoli, Barbara ; Lusso, Paolo ; Malnati, Mauro S. / Calibrated real-time PCR assay for quantitation of human herpesvirus 8 DNA in biological fluids. In: Journal of Clinical Microbiology. 2002 ; Vol. 40, No. 12. pp. 4652-4658.
@article{ed353953bdbf4ad5879d85c5fea2f211,
title = "Calibrated real-time PCR assay for quantitation of human herpesvirus 8 DNA in biological fluids",
abstract = "Accurate laboratory tests for the diagnosis of active human herpesvirus 8 (HHV-8) infection are becoming essential to study the pathogenesis of HHV-8-associated tumors and for the clinical management of HHV-8-infected individuals. We have developed a highly sensitive, calibrated quantitative real-time PCR assay for the measurement of cell-free HHV-8 DNA in body fluids, based on the addition of a synthetic DNA calibrator prior to DNA extraction. The calibrator controls each sample for the presence of PCR inhibitors, determines a cutoff value of sensitivity for negative samples, and normalizes positive samples for the efficiency of DNA recovery. The assay shows a wide dynamic range of detection (between 1 and 106 viral genome equivalents/reaction) and a high degree of accuracy even in the presence of high amounts (up to 1 μg) of human genomic DNA. Moreover, the assay has a very high sensitivity (lower detection limit, 10 genome equivalents/ml) and a high degree of reproducibility and repeatability with a coefficient of variation (CV) of",
author = "Francesco Broccolo and Giuseppe Locatelli and Loredana Sarmati and Sara Piergiovanni and Fabrizio Veglia and Massimo Andreoni and Stefano Butt{\`o} and Barbara Ensoli and Paolo Lusso and Malnati, {Mauro S.}",
year = "2002",
month = "12",
day = "1",
doi = "10.1128/JCM.40.12.4652-4658.2002",
language = "English",
volume = "40",
pages = "4652--4658",
journal = "Journal of Clinical Microbiology",
issn = "0095-1137",
publisher = "American Society for Microbiology",
number = "12",

}

TY - JOUR

T1 - Calibrated real-time PCR assay for quantitation of human herpesvirus 8 DNA in biological fluids

AU - Broccolo, Francesco

AU - Locatelli, Giuseppe

AU - Sarmati, Loredana

AU - Piergiovanni, Sara

AU - Veglia, Fabrizio

AU - Andreoni, Massimo

AU - Buttò, Stefano

AU - Ensoli, Barbara

AU - Lusso, Paolo

AU - Malnati, Mauro S.

PY - 2002/12/1

Y1 - 2002/12/1

N2 - Accurate laboratory tests for the diagnosis of active human herpesvirus 8 (HHV-8) infection are becoming essential to study the pathogenesis of HHV-8-associated tumors and for the clinical management of HHV-8-infected individuals. We have developed a highly sensitive, calibrated quantitative real-time PCR assay for the measurement of cell-free HHV-8 DNA in body fluids, based on the addition of a synthetic DNA calibrator prior to DNA extraction. The calibrator controls each sample for the presence of PCR inhibitors, determines a cutoff value of sensitivity for negative samples, and normalizes positive samples for the efficiency of DNA recovery. The assay shows a wide dynamic range of detection (between 1 and 106 viral genome equivalents/reaction) and a high degree of accuracy even in the presence of high amounts (up to 1 μg) of human genomic DNA. Moreover, the assay has a very high sensitivity (lower detection limit, 10 genome equivalents/ml) and a high degree of reproducibility and repeatability with a coefficient of variation (CV) of

AB - Accurate laboratory tests for the diagnosis of active human herpesvirus 8 (HHV-8) infection are becoming essential to study the pathogenesis of HHV-8-associated tumors and for the clinical management of HHV-8-infected individuals. We have developed a highly sensitive, calibrated quantitative real-time PCR assay for the measurement of cell-free HHV-8 DNA in body fluids, based on the addition of a synthetic DNA calibrator prior to DNA extraction. The calibrator controls each sample for the presence of PCR inhibitors, determines a cutoff value of sensitivity for negative samples, and normalizes positive samples for the efficiency of DNA recovery. The assay shows a wide dynamic range of detection (between 1 and 106 viral genome equivalents/reaction) and a high degree of accuracy even in the presence of high amounts (up to 1 μg) of human genomic DNA. Moreover, the assay has a very high sensitivity (lower detection limit, 10 genome equivalents/ml) and a high degree of reproducibility and repeatability with a coefficient of variation (CV) of

UR - http://www.scopus.com/inward/record.url?scp=18744393373&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=18744393373&partnerID=8YFLogxK

U2 - 10.1128/JCM.40.12.4652-4658.2002

DO - 10.1128/JCM.40.12.4652-4658.2002

M3 - Article

VL - 40

SP - 4652

EP - 4658

JO - Journal of Clinical Microbiology

JF - Journal of Clinical Microbiology

SN - 0095-1137

IS - 12

ER -