Development of a Multiplex HIV/TB Diagnostic Assay Based on the Microarray Technology

Abstract

Currently there are diagnostic tests available for human immunodeficiency virus (HIV) and tuberculosis (TB); however, they are still diagnosed separately, which can delay treatment in cases of co-infection. Here we report on a multiplex microarray technology for the detection of HIV and TB antibodies using p24 as well as TB CFP10, ESAT6 and pstS1 antigens on epoxy-silane slides. To test this technology for antigen–antibody interactions, immobilized antigens were exposed to human sera spiked with physiological concentrations of primary antibodies, followed by secondary antibodies conjugated to a fluorescent reporter. HIV and TB antibodies were captured with no cross-reactivity observed. The sensitivity of the slides was compared to that of high-binding plates. We found that the slides were more sensitive, with the detection limit being 0.000954 µg/mL compared to 4.637 µg/mL for the plates. Furthermore, stability studies revealed that the immobilized antigens could be stored dry for at least 90 days and remained stable across all pH and temperatures assessed, with pH 7.4 and 25 °C being optimal. The data collectively suggested that the HIV/TB multiplex detection technology we developed has the potential for use to diagnose HIV and TB co-infection, and thus can be developed further for the purpose.

Keywords

• HIV-1 p24
• <i>M.tb</i> CFP10
• <i>M.tb</i> ESAT6
• <i>M.tb</i> pstS1
• multiplex microarray
• diagnosis