Zaheer Parpia

p24 antigen rapid test for diagnosis of acute pediatric HIV infection.

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.

Membrane-based plasma collection device for point-of-care diagnosis of HIV

A major requirement for the development of point-of-care tests for the detection of disease analytes is the need to separate plasma from whole blood in an efficient and rapid manner. Furthermore, the separated plasma must be able to elute efficiently the analyte of interest and serve effectively as a physical matrix to deliver the equivalent of neat plasma for downstream diagnostic analysis. Additionally, many applications require the use of heat shock to liberate immunocomplexed antigen found in the collected plasma. A membrane-based filter method is reported for rapid and efficient collection of plasma from a whole blood sample that is compatible with heat shock. Using pediatric human immunodeficiency virus as an example, this device elutes 100% of the input p24 core antigen post-collection and enables heat shock of plasma samples identical to neat plasma treatment.

A lateral flow-based ultra-sensitive p24 HIV assay utilizing fluorescent microparticles.

The current demand for HIV diagnostic tests in resource-limited settings is not being fully addressed by the currently available nucleic acid test or enzyme-linked immunosorbent assay-based p24 antigen assays. This is primarily due to their high cost, their requirement for controlled laboratory conditions and/or personnel, or the relatively long turn-around time for test results. Self performing lateral flow assays address all of these issues but to date have not reached the level of analytical sensitivity for HIV p24 demonstrated by current generation enzyme-linked immunosorbent assay technology. This report presents an initial prototype lateral flow assay for HIV p24 antigen capable of achieving subpicogram per milliliter limits of analytical sensitivity and requiring less than 40 minutes to yield results.