Toward Development of a Label-Free Detection Technique for Microfluidic Fluorometric Peptide-Based Biosensor Systems

Nikita Sitkov; Tatiana Zimina; Alexander Kolobov; Vladimir Karasev; Alexander Romanov; Viktor Luchinin; Dmitry Kaplun

doi:10.3390/mi12060691

Toward Development of a Label-Free Detection Technique for Microfluidic Fluorometric Peptide-Based Biosensor Systems

Micromachines (Basel). 2021 Jun 13;12(6):691. doi: 10.3390/mi12060691.

Authors

Nikita Sitkov¹, Tatiana Zimina¹, Alexander Kolobov², Vladimir Karasev¹, Alexander Romanov¹, Viktor Luchinin¹, Dmitry Kaplun³

Affiliations

¹ Department of Micro- and Nanoelectronics, Saint Petersburg Electrotechnical University "LETI", 197376 Saint Petersburg, Russia.
² Institute of Highly Pure Biopreparations, 197110 Saint Petersburg, Russia.
³ Department of Automation and Control Processes, Saint Petersburg Electrotechnical University "LETI", 197376 Saint Petersburg, Russia.

Abstract

The problems of chronic or noncommunicable diseases (NCD) that now kill around 40 million people each year require multiparametric combinatorial diagnostics for the selection of effective treatment tactics. This could be implemented using the biosensor principle based on peptide aptamers for spatial recognition of corresponding protein markers of diseases in biological fluids. In this paper, a low-cost label-free principle of biomarker detection using a biosensor system based on fluorometric registration of the target proteins bound to peptide aptamers was investigated. The main detection principle considered includes the re-emission of the natural fluorescence of selectively bound protein markers into a longer-wavelength radiation easily detectable by common charge-coupled devices (CCD) using a specific luminophore. Implementation of this type of detection system demands the reduction of all types of stray light and background fluorescence of construction materials and aptamers. The latter was achieved by careful selection of materials and design of peptide aptamers with substituted aromatic amino acid residues and considering troponin T, troponin I, and bovine serum albumin as an example. The peptide aptamers for troponin T were designed in silico using the «Protein 3D» (SPB ETU, St. Petersburg, Russia) software. The luminophore was selected from the line of ZnS-based solid-state compounds. The test microfluidic system was arranged as a flow through a massive of four working chambers for immobilization of peptide aptamers, coupled with the optical detection system, based on thick film technology. The planar optical setup of the biosensor registration system was arranged as an excitation-emission cascade including 280 nm ultraviolet (UV) light-emitting diode (LED), polypropylene (PP) UV transparent film, proteins layer, glass filter, luminophore layer, and CCD sensor. A laboratory sample has been created.

Keywords: lab-on-a-chip; label-free detection; microfluidic biochip; peptide aptamers; peptide biosensor; protein biomarkers; troponin T.

Grants and funding

19-32-90020/Russian Foundation for Basic Research