Development and validation of an instrument-free system for DNA target detection based on the ultra-monodisperse gold nanospheres

Objective: to develop a simple and robust technology for DNA targets detection for the diagnosis of socially significant diseases, based on a colorimetric aggregation test using gold nanoparticles. Material and methods. Gold nanospheres were synthesized using liquid-phase chemistry methods. The physicochemical parameters of the nanoparticles were controlled by absorption spectroscopy, dynamic light scattering, and electron microscopy. The rational design of the specific sequences of target genomic fragments and oligonucleotides included in the test system, was carried out taking into account the thermodynamic constants and using genomic databases. The analytical validation of the test system was carried out using a model for detecting the DNA of Babesia species, the causative agent of human and animal pyroplasmosis, by cross-comparing the characteristics with a polymerase chain reaction with electrophoretic results detection and in a “real-time” format. Results. A pilot test system has been developed for the specific detection of up to 10 ng of DNA in blood samples by visual discrimination of the color of the reaction mixture, within analysis duration about 30 min. The size effect of nanoparticles in the range from 15 to 60 nm on the minimum detection limit of DNA targets has been revealed. Conclusion. This paper presents the results of the development and analytical validation of a pilot version of an express test system for DNA diagnostics based on gold nanoparticle tags and visual color discrimination of the reaction mixture.