Real-time immuno-PCR assay for detecting PCBs in soil samples

Abstract

A fast and robust assay, based on immuno-polymerase chain reaction (IPCR) techniques, was developed for the detection of polychlorinated biphenyls (PCBs) in soil samples. Real-time IPCR (rt-IPCR) is a powerful technique that combines enzyme-linked immunosorbent assay (ELISA) with the specificity and sensitivity of PCR. In our assay, indirect ELISAs based on immobilization of PCB37 hapten-ovalbumin conjugates was used for evaluation of the immune response. The effect of optimal reagent concentrations to reduce background fluorescence was also investigated. Using the optimized assay, the linear sensitivity range of the assay covered more than six orders of magnitude, and the minimum detection limits reached 5 fg ml(-1) antigen. Rt-IPCR was tested for its cross-reactivity profiles using four selected congeners and four Aroclor products. The assays were highly specific for congeners but less specific for Aroclor1242. We took four soil samples to validate the method, and the results were confirmed by gas chromatography/mass spectrometry (GC/MS). The rt-IPCR results for soil samples correlated well with the concentrations of PCBs obtained by GC/MS (r = 0.99, n = 6). These data indicate that this highly specific, sensitive, and robust assay can be modified for detecting PCB compounds in the environment.

Novel methodologies in analysis of small molecule biomarkers and living cells

“…Their study revealed that the real-time immuno-PCR method is highly sensitive and precise; therefore, it is a promising tool in the trace analysis of environmental endocrine disruptors [11]. Their group also developed real-time immuno-PCR to detect toxicological and carcinogenic polychlorinated biphenyls in environmental samples [12]. All of their results were confirmed by conventional ELISA methods and further validated the reliability of quantitative immuno-PCR.…”

DNA for Non-nucleic Acid Sensing

“…The possibility of labelling the secondary antibody with a nucleotide sequence and to implement sequence amplification using PCR has been explored (Sano et al 1992). This approach improves the sensitivity of ELISA tests by a factor of 100-10,000 and has been widely used for the detection of rare infectious diseases (Guo et al 2006), toxins in food (Kuczius et al 2009;Zhang et al 2010) or pollutants in aqueous media (Zhuang and Zhou 2009;Chen and Zhuang 2009). However, PCR requires the implementation of complex protocols that need special instruments and qualified personnel.…”

Immunoanalysis Methods for the Detection of Dioxins and Related Chemicals

“…Recently, it has been introduced to detect environmental contaminants, such as dioxin. For example, Chen et al [118] first developed a fluorescent quantitative IPCR and assessed its potential for the detection of antibodies recognizing PCBs in soil samples, in which the PCB concentrations were as low as 10 fg·mL −1 . Recovery rates were 95.0–105.0% and the Rt-IPCR results correlated well with the concentration of PCBs obtained by GC/MS (r = 0.99, n = 6).…”