Real-time quantitative PCR for human herpesvirus 6 DNA

G. Locatelli, F. Santoro, F. Veglia, A. Gobbi, P. Lusso, M. S. Malnati

Research output: Contribution to journalArticlepeer-review

Abstract

The diagnosis of human herpesvirus 6 (HHV-6) infection represents a complex issue because the most widely used diagnostic tools, such as immunoglobulin G antibody titer determination and qualitative DNA PCR with blood cells, are unable to distinguish between latent (clinically silent) and active (often clinically relevant) infection. We have developed a new, highly sensitive, quantitative PCR assay for the accurate measurement of HHV-6 DNA in tissue-derived cell suspensions and body fluids. The test uses a 5′ nuclease, fluorogenic assay combined with real-time detection of PCR amplification products with the ABI PRISM 7700 sequence detector system. The sensitivity of this method is equal to the sensitivity of a nested PCR protocol (lower detection limit, 1 viral genome equivalent/test) for both the A and the B HHV-6 subgroups and shows a wider dynamic range of detection (from 1 to 106 viral genome equivalents/test) and a higher degree of accuracy, repeatability, and reproducibility compared to those of a standard quantitative-competitive PCR assay developed with the same reference DNA molecule. The novel technique is versatile, showing the same sensitivity and dynamic range with viral DNA extracted from different fluids (i.e., culture medium or plasma) or from tissue-derived cell suspensions. Furthermore, by virtue of its high-throughput format, this method is well suited for large epidemiological surveys.

Original languageEnglish
Pages (from-to)4042-4048
Number of pages7
JournalJournal of Clinical Microbiology
Volume38
Issue number11
Publication statusPublished - 2000

ASJC Scopus subject areas

  • Microbiology (medical)
  • Microbiology

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