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 user-friendly technology for the detection of DNA targets for the diagnosis of socially significant diseases based on a colorimetric aggregation assay using gold nanoparticles. Materials and Methods: Gold nanospheres were synthesized by liquid-phase chemistry methods. The physicochemical parameters of the nanoparticles were controlled using absorption spectroscopy, dynamic light scattering, and electron microscopy. Rational design of specific genomic target sequences and oligonucleotides included in the test system was carried out considering thermodynamic constants and using genomic databases. Analytical validation of the test system was performed using a model for the detection of Babesia species DNA — the causative agent of piroplasmosis in humans and animals — by cross-comparison of characteristics with polymerase chain reaction (PCR) with electrophoretic detection in real-time format. Results: A pilot version of the test system for specific detection of up to 10 ng of DNA in blood samples was developed, based on visual discrimination of the color of the reaction mixture with an analysis duration of approximately 30 minutes. The size effect of nanoparticles in the range from 15 to 60 nm on the minimum detection limit of DNA targets was established. Conclusion: This study presents the results of the development and analytical validation of a pilot express test system for DNA diagnostics based on gold nanoparticle labeling and visual color discrimination of the reaction mixture.