Background:  Over the past decade, certain nanotechnology-based techniques have been widely evaluated in medical testing and could provide new tools for clinical diagnosis due to their potential for high degrees of sensitivity, high specificity, multiplexing capabilities and ability to operate without enzymes. In 2007, we described a gold nanoparticle-based bio-barcode amplification (BCA) assay for early and sensitive detection of HIV-1 capsid (p24) antigen. Now we present a simple and sensitive immunoassay using highly fluorescent europium nanoparticles for detection of HIV-1 p24.

Method:  The primary anti-p24 antibody coated-microplate was used to capture the target (p24 antigen). Biotin-labeled secondary anti-p24 antibody and streptavidin-labeled, highly fluorescent europium\nanoparticles and streptavidin-europium⁺ chelates were used for signal amplification. Fluorescence signals can be measured immediately after the reaction and the entire assay can be completed within 2 to 3 hours.

Results:  Our preliminary results indicate that the lower limit of detection achievable by this method is 0.5 pg/mL, which is significantly more sensitive than conventional enzyme-linked immunosorbent assay (ELISA). Although our ultrasensitive BCA assay can detect 0.1 pg/ml of HIV-1 p24 and was approximately 150-fold more sensitive than ELISA, the europium particle-based immunoassay is simpler and quicker and does not need expensive equipment for detection. The evaluation of the sensitivity and specificity of the assay is ongoing.

Conclusions:  Our results indicate that nanoparticle-based techniques can dramatically improve the sensitivity of HIV-1 p24 antigen detection in the absence of enzymatic reactions. The preliminary evaluation of the method based on the small samples indicates that the HIV-1 p24 antigen BCA may provide a new tool for sensitive and early detection of HIV-1 p24 antigen in settings where HIV-1 RNA testing is currently not routinely performed. europium⁺ nanoparticle-based immunoassay could provide a rapid and sensitive tool for detection of various pathogen markers. It may be suitable for further application in high-throughput screening and point-of-care use and potentially in resource limited settings. It can also be expanded to other pathogens since key reagents used in the method are universal.