p24 Antigen Rapid Test for Diagnosis of Acute Pediatric HIV Infection

Zaheer A. Parpia, PhD; Robert Elghanian, PhD; Arman Nabatiyan, PhD; Diana R. Hardie, MD; David M. Kelso, PhD

Disclosures

J Acquir Immune Defic Syndr. 2010;55(4):413-419. 

In This Article

Abstract and Introduction

Abstract

Currently, the majority of HIV-infected infants are found within limited-resource settings, where inadequate screening for HIV due to the lack of access to simple and affordable point-of-care tests impedes implementation of antiretroviral therapy. Here we report development of a low-cost dipstick p24 antigen assay using a visual readout format that can facilitate the diagnosis of HIV for infants in resource-poor conditions. A heat shock methodology was developed to optimize disruption of immune complexes present in the plasma of infected infants. The analytical sensitivity of the assay using recombinant p24 antigen is 50 pg/mL (2 pM) with whole virus detection as low as 42.5k RNA copies per milliliter plasma. In a blinded study comprising 51 archived infant samples from the Women and Infants Transmission Study, our assay demonstrated an overall sensitivity and specificity of 90% and 100%, respectively. In field evaluations of 389 fresh samples from South African infants, a sensitivity of 95% and specificity of 99% was achieved. The assay is simple to perform, requires minimal plasma volume (25 μL), and yields a result in less than 40 minutes making it ideal for implementation in resource-limited settings.

Introduction

Approximately 1.5 million infants are born to HIV-infected women each year,[1] majority of whom are not tested for HIV until it is too late for optimal antiretroviral therapy (ART). Without treatment, the mortality rate in HIV-infected infants can be as high as 45% by the first birthday and 59% by the second.[2] Recent studies have demonstrated that early HIV diagnosis and prompt ART intervention can reduce infant mortality by 76% and HIV progression by 75%.[3] Such studies have contributed to a change in treatment guidelines by the World Health Organization to initiate ART therapy in infants as soon as they are diagnosed with HIV.[1]

Worldwide, there is vast disparity in health care provisions for HIV-infected infants in the developed world and those in resource-poor countries. In resource-limited countries, where 90% of the exposed infants are found, several obstacles such as limited screening programs for HIV and the lack of a simple and affordable point-of-care diagnostic currently impede the widespread implementation of ARTs. The current gold standard for HIV testing, DNA polymerase chain reaction (PCR), is not suited for implementation in these settings because of the long turn-around-times and inefficiencies involved in transporting samples to central laboratories and returning results to clinics. These inefficiencies lead to poor follow-up and low turn-outs for testing. Rapid antibody tests make diagnostic results available on the same visit, but cannot be used to diagnose infection as HIV exposed infants can retain maternal antibodies for up to 18 months.[4]

Various studies have highlighted the utility of HIV core p24 antigen detection for adult and pediatric screening,[5,6] prediction of disease progression,[7,8] and monitoring the effectiveness of ART.[9,10] An excellent overview of the work has been presented by Schupbach.[11] However, these studies have been carried out with enzyme-linked immunosorbent assay-based systems which are similar in complexity to PCR techniques in that they are time intensive and require expensive laboratory equipment and technical expertise which are not readily available in most low-income and middle-income countries.

Here, we report the development of a rapid p24 antigen dipstick assay designed to diagnose HIV in infant plasma samples with minimal operator input. The assay uses a visual carbon-based reporter label functionalized with antibodies against HIV p24 antigen, and a heat-shock pretreatment step was incorporated into the assay to disrupt immune complexes. The analytical sensitivity of our assay was determined to be in the single picomolar range in model systems of recombinant p24 antigen and intact HIV virus spiked in human plasma. We subsequently assessed the diagnostic performance of our p24 antigen assay for HIV detection in a set of 51 archived infant samples obtained from the US Women and Infants Transmission Study (WITS) and with 389 target infant specimens from a screening laboratory in South Africa. The sensitivity (>90%) and specificity (>99%) measurements resulting from testing infant samples from United States and Sub-Saharan Africa indicates that our prototype assay is suitable for further development to meet infant diagnostic needs in affected countries.

Comments

3090D553-9492-4563-8681-AD288FA52ACE
Comments on Medscape are moderated and should be professional in tone and on topic. You must declare any conflicts of interest related to your comments and responses. Please see our Commenting Guide for further information. We reserve the right to remove posts at our sole discretion.
Post as:

processing....