PCR-amplified Immunoassay (Immuno-PCR): Principle of the Method, Variants of Execution, Possibilities and Prospects of Use for the Detection of Pathogenic Biological Agents

Abstract

Enzyme immunoassay and polymerase chain reaction have become the «gold standard» for the detection of biological pathogens. The method of amplified immunoassay – immuno-PCR allows to combine both methods into a single platform to preserve their advantages and to achieve high sensitivity of the analysis. The purpose of this work is to consider the possibilities and prospects of using PCR-amplified immunoassay for the detection of pathogenic biological agents. Immuno-PCR makes it possible to detect various non-nucleic antigenic determinants in PCR by amplifying a DNA tag conjugated with a specific antibody. The registration of the results is also possible in real time as in the real-time PCR test systems. The main methodological issues in the immuno-PCR technology are: the choice of a carrier of biomolecule complexes, the choice of a method for conjugation of detection antibodies and a reporter nucleic acid, optimization of methods for amplifying signal DNA and accounting for results, and development of methods for reducing background indicators. We consider it necessary to carry out research and development work on the development and the creation of diagnostic kits based on immuno-PCR. With regard to the task of detecting small and trace amounts of antigens of pathogenic biological agents, the most likely diagnostic «niche» of the immuno-PCR method will be the detection of toxins of microbial and non-microbial origin, the minimum clinically significant dose for which is less than the sensitivity of the corresponding immunochemical test systems. Taking into account the prospects for the development of the method, in future it is possible to develop such test systems for the detection of hapten analytes, for example, some toxicants of non-biological origin