RESUMEN
A method for detection of cancer-associated protein D-NFATc1 in serum using nanowire (NW) biosensor based on field-effect nanotransistor is developed. Field-effect nanotransistor was fabricated on the basis of «silicon-on-insulator¼ structures. For the biospecific detection of target protein, the NW surface was modified with aptamers against the target protein. Using the 3 um-NW enabled to obtain stable source-drain characteristics and to register D-NFATc1 in serum at concentration of 2.5 x 1014 M in the mode of drain-source current vs. gate voltage characteristics measurements. Data collection in the mode of drain-source current vs. gate voltage characteristics measurements was carried out with the use of high-speed data collection system running TURBO NBS software.
Asunto(s)
Técnicas Biosensibles , Factores de Transcripción NFATC/sangre , Nanocables , Proteínas de Neoplasias/sangre , Programas Informáticos , Transistores Electrónicos , Técnicas Biosensibles/instrumentación , Técnicas Biosensibles/métodos , Humanos , Sensibilidad y EspecificidadRESUMEN
The nanowire (NW) detection is one of fast-acting and high-sensitive methods allowing to reveal potentially relevant protein molecules. A NW biosensor based on the silicon-on-insulator (SOI)-structures was used for biospecific label-free detection of NFAT 1 (D-NFAT 1) oncomarker in real time. For this purpose, SOI-nanowires (NWs) were modified with aptamers against NFAT 1 used as molecular probes. It was shown that using this biosensor it is possible to reach the sensitivity of ~10(-15) M. This sensitivity was comparable with that of the NW biosensor with immobilized antibodies used as macromolecular probes. The results demonstrate promising approaches used to form the sensor elements for high-sensitive disease diagnostics.
Asunto(s)
Aptámeros de Nucleótidos/química , Técnicas Biosensibles/instrumentación , Factores de Transcripción NFATC/análisis , Nanocables/química , Proteínas de Neoplasias/análisis , Anticuerpos Monoclonales/química , Aptámeros de Nucleótidos/síntesis química , Técnicas Biosensibles/economía , Técnicas Electroquímicas , Humanos , Límite de Detección , Dióxido de Silicio/química , Soluciones , Succinimidas/químicaRESUMEN
Achievement of the concentration detection limit for proteins at the level of the reverse Avogadro number determines the modern development of proteomics. In this review, the possibility of approximating the reverse Avogadro number by using nanotechnological methods (AFM-based fishing with mechanical and electrical stimulation, nanowire detectors, and other methods) are discussed. The ability of AFM to detect, count, visualize and characterize physico-chemical properties of proteins at concentrations up to 10(-17)-10(-18) M is demonstrated. The combination of AFM-fishing with mass-spectrometry allows the identification of proteins not only in pure solutions, but also in multi-component biological fluids (serum). The possibilities to improve the biospecific fishing efficiency by use of SOMAmers in both AFM and nanowire systems are discussed. The paper also provides criteria for evaluation of the sensitivity of fishing-based detection systems. The fishing efficiency depending on the detection system parameters is estimated. The practical implementation of protein fishing depending on the ratio of the sample solution volume and the surface of the detection system is discussed. The advantages and disadvantages of today's promising nanotechnological protein detection methods implemented on the basis of these schemes.
Asunto(s)
Microscopía de Fuerza Atómica/métodos , Proteínas/análisis , Animales , Técnicas Biosensibles/métodos , Técnicas Electroquímicas/instrumentación , Técnicas Electroquímicas/métodos , Ensayo de Inmunoadsorción Enzimática , Humanos , Límite de Detección , Espectrometría de Masas , Microscopía de Fuerza Atómica/instrumentación , Técnicas de Sonda Molecular , Nanotecnología/métodos , Nanocables , Proteínas/metabolismo , Resonancia por Plasmón de Superficie/métodosRESUMEN
The atomic-force microscopy-based method of irreversible chemical AFM-fishing (AFM-IF(Ch)) has been developed for the detection of proteins at ultra-low concentrations in solution. Using this method, a very low concentration of horseradish peroxidase (HRP) protein (10(-17) M) was detected in solution. A theoretical model that allows the description of obtained experimental data, is proposed. This model takes into consideration both the transport of the protein from the bulk solution onto the AFM-chip surface and its irreversible binding to the activated area.
