A bioorganometallic approach for rapid electrochemical analysis of human immunodeficiency virus type-1 reverse transcriptase in serum

Highlights: → Two-dimensional electrochemical assay of HIV-1 reverse transcriptase in serum is presented. → XPS and ToF-SIMS confirmed binding of synthetic ferrocene-lipoic acid to gold. → Dynamic range was 100-500 pg mL-1 with an LOD of 50 pg mL-1. → The biosensor allows sensitive and selective detection of reverse transcriptase in 20 s. - Abstract: Determination of low-abundance human immunodeficiency virus (HIV) enzymes is essential for early detection of the viral infection. Designed for the rapid detection of the virus in human serum, a rapid and ultrasensitive electrochemical assay for HIV-1 reverse transcriptase (HIV-1 RT) is presented in this article. The assay format is based on the formation of a self-assembled monolayer (SAM) of a synthesized ferrocene (Fc)-labeled lipoic acid onto a gold electrode. X-ray photoelectron spectroscopy (XPS) and Time-of-Flight secondary ion mass spectrometry (ToF-SIMS) were employed to confirm the binding of the Fc-labeled lipoic acid to the gold via the Au-S bond. A short RT-specific peptide, VEAIIRILQQLLFIH, was covalently attached to the Fc-labeled lipoic acid. Square wave voltammetry (SWV) offered a two-dimensional measurement of RT based on the anodic shift and reduction of current density of the Fc redox signal upon binding of RT to its specific peptide. This allowed a linear quantification of the target RT in the range of 100-500 pg mL-1, equivalent to 85.5-427.4 fM, with a detection limit of 50 pg mL-1 (42.7 fM). The developed biosensor is inexpensive, easy to prepare and operate, and allows a highly selective detection of RT in 20 s.