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

Sally M. McFall; Robin L. Wagner; Sujit R. Jangam; Douglas H. Yamada; Diana Hardie; David M. Kelso

doi:10.1016/j.jviromet.2015.01.005

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

Sally M. McFall^*, Robin L. Wagner, Sujit R. Jangam, Douglas H. Yamada, Diana Hardie, David M. Kelso

^*Corresponding author for this work

Biomedical Engineering

Research output: Contribution to journal › Article › peer-review

48 Scopus citations

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.

Original language	English (US)
Pages (from-to)	37-42
Number of pages	6
Journal	Journal of Virological Methods
Volume	214
DOIs	https://doi.org/10.1016/j.jviromet.2015.01.005
State	Published - Mar 5 2015

Funding

This study was supported by the Bill and Melinda Gates Foundation Grand Challenges in Global Health grant 37774 . Real-time PCR reagents and advice were provided by Abbott Molecular Inc. Des Plaines, IL. 8E5-LAV cells were provided by the Virology Quality Assurance Laboratory, Rush Presbyterian; St. Luke's Medical Center. Adult HIV-1 negative blood was provided by Core Lab, NorthShore University HealthSystems, Evanston, IL; and infant blood was provided by Children's Memorial Hospital, Chicago, IL. Thanks to Stuart Knowles for help with data analysis, and to Mark Fisher for photography assistance.

Keywords

Early infant diagnosis
HIV
Polymerase chain reaction
Provirus

ASJC Scopus subject areas

Virology

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

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10.1016/j.jviromet.2015.01.005

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title = "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",

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.",

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AU - Wagner, Robin L.

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AU - Yamada, Douglas H.

AU - Hardie, Diana

AU - Kelso, David M.

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AB - 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.

KW - Early infant diagnosis

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KW - Polymerase chain reaction

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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

Abstract

Funding

Keywords

ASJC Scopus subject areas

UN SDGs

Access to Document

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