Direct quantification of HIV-1 RNA by capillary electrophoresis with laser-induced fluorescence

Abstract

HIV-1 RNA was quantitated directly by capillary electrophoresis with laser-induced fluorescence (CE-LIF). CE-LIF was used to analyze cellular RNA and various nucleotide complexes. A fluorescently labeled DNA probe (DNA/RNA complex) in conjunction with thiazole orange intercalator was determined to have optimal stability and sensitivity for RNA analysis. Based on this observation, a hybridization method using a HIV-specific fluorescently labeled probe with analysis by CE-LIF was developed. Plasma samples from a HIV-seropositive patient were lysed to obtain RNA, hybridized with the HIV-specific probe and analyzed by CE-LIF. As little as 19 fg (1710 copies per 1 ml of starting plasma) of HIV RNA can be reliably and quantitatively detected. CE-LIF appears to be an efficient and sensitive method to quantitatively analyze RNA from a variety of sources.