Elsevier

Journal of Virological Methods

Volume 214, 15 March 2015, Pages 37-42
Journal of Virological Methods

A simple and rapid DNA extraction method from whole blood for highly sensitive detection and quantitation of HIV-1 proviral DNA by real-time PCR

https://doi.org/10.1016/j.jviromet.2015.01.005Get rights and content
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open access

Highlights

  • This FINA method with internal control (IC) was developed to detect proviral HIV-1 DNA from whole blood samples using a modification of Abbott's HIV-1 RealTime Assay for early infant diagnosis.

  • The limit of quantification is 10 copies/100 μl whole blood and as little as a single proviral copy can be detected.

  • In a preliminary field test with 61 South African infant specimens, the FINA assay demonstrated 100% sensitivity and specificity.

Abstract

Early diagnosis and access to treatment for infants with human immunodeficiency virus-1 (HIV-1) is critical to reduce infant mortality. The lack of simple point-of-care tests impedes the timely initiation of antiretroviral therapy. The development of FINA, filtration isolation of nucleic acids, a novel DNA extraction method that can be performed by clinic personnel in less than 2 min has been reported previously. In this report, significant improvements in the DNA extraction and amplification methods are detailed that allow sensitive quantitation of as little as 10 copies of HIV-1 proviral DNA and detection of 3 copies extracted from 100 μl of whole blood. An internal control to detect PCR inhibition was also incorporated. In a preliminary field evaluation of 61 South African infants, the FINA test demonstrated 100% sensitivity and specificity. The proviral copy number of the infant specimens was quantified, and it was established that 100 microliters of whole blood is required for sensitive diagnosis of infants.

Keywords

HIV
Early infant diagnosis
Provirus
Polymerase chain reaction

Abbreviations

DBS
dried blood spot
FINA
filtration isolation of nucleic acids
POC
point-of-care
LOD
limit of detection
LOQ
limit of quantitation
HPR
hydroxypyruvate reductase
IC
internal control

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