Search

[Display omitted] • The VHH displaying phage fabricated for VEGF detection. • The preparation and operation simplified by labeling steps elimination. • The results revealed excellent specificity and reliability of proposed immunosensor. • Analysis of results showed valuable LOD, LOQ and wide linear range. Herein the pioneering label-free electrochemistry-based immunosensor with exceptional sensitivity for the rapid and efficient detection of vascular endothelial growth factor (VEGF) was precisely designed based on the phage display technique. The specific variable domain of a heavy-chain antibody (VHH)-displaying phage against VEGF 8–109 was successfully immobilized on Au nanoparticles deposited gold electrode modified by 3-mercapto propionic acid and 11-mercaptoundecanoic acid and then, carbodiimide coupling reaction using N-hydroxysuccinimide and N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride. Electrochemical characterizations of the proposed immunosensor were performed by PM-FT-IRRAS, scanning electron microscopy, cyclic voltammetry and electrochemical impedance spectroscopy techniques. Electrochemical measurements were performed using differential pulse voltammetry in phosphate buffer containing [Fe(CN) 6 ]3–/4– ions as a probe, where the decline in the peak current value was related to a concentration of the antigen cast on the surface of the immunosensor. Under the optimum conditions (pH 7.4, interval time 20 s, and step potential 0.1 V), a linear range of response for the detection of VEGF 8–109 by immunosensor was 0.03–10,000 pg/mL (in log concentration), with a limit of detection of 11 fg/mL. The specificity of the immunosensor was also investigated against some typical blood growth factors. The novel immunosensor was applied to evaluate VEGF levels in human serum specimens clinically. The recommended immunosensor was uncomplicated, reasonably priced, and showed adequate accuracy and specificity, which could be applied in the early diagnosis of tumors. [ABSTRACT FROM AUTHOR]

• Fabrication of novel label free chemically efficient electrochemical immunosensor for TNF-α direct determination. • Using FFT admittance signal for TNF-α concentration monitoring as advanced modern electroanalytical technique. • Preparing effective imbed for TNF-α antibody immobilizing using HKUST-1/Au NPs. • Reaching lower detection limit and higher sensitivity compared with conventional quantitative electrochemical methods. A new simple label-free electrochemical immunosensor was designed for sensitive and selective determination of tumor necrosis factor-alpha (TNF-α), through gold nanoparticles (GNPs) incorporated copper benzene tricarboxylate metal organic frameworks (HKUST-1) modified gold electrode, where TNF-α specific monoclonal antibody was bonded to the electrode surface through shaped self-assembled monolayer (SAM) of mercapto compounds. The fabrication steps were monitored by field emission scanning electron microscopy (FE-SEM), cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). Under optimized experimental conditions, fast Fourier transform admittance voltammetry (FFTADV) used for TNF-α measurement. A wide linear range of current response towards the TNF-α concentration from 0.0005 to 50 ng/mL and LOD value of 0.15 pg/mL were achieved using FFTADV. The described immunosensor had adequate sensitivity and selectivity towards direct determination of TNF-α in complex biological matrix. [ABSTRACT FROM AUTHOR]

A novel label-free electrochemical immunosensor was fabricated for selective and sensitive detection of Candidatus Phytoplasma Aurantifolia (CPA), the main factor of witches broom diseases, in tree sap samples. CPA antibody was immobilized on a thiol terminated surface of a gold electrode modified by gold nanoparticles; the surface was then activated by mixture of N -(3-dimethylaminopropyl)- N -ethylcarbodiimide hydrochloride (EDC) and N -HydroxySuccinamide (NHS). Scanning electron microscopy (SEM), cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were employed for surface characterization. All the electrochemical measurements of CPA were accomplished based on the [Fe(CN) 6 ]3−/4− solution response as the standard redox probe. CPA detection was carried out using square wave voltammetry (SWV) technique, at frequency of 20 Hz and amplitude of 0.1 V. In order to reach the maximum efficiency in performance of the immunosensor, Anti-CPA 1 mg mL−1, pH 7.4 and the incubation time of 45 min were selected as optimized experimental conditions. According to the calibration curve data, the obtained linear range was 0.005 to 100 μg mL−1, with limit of detection (LOD) of 1.5 ng mL−1, which was lower than the value reported for ELISA method applied for CPA detection. The immunosensor demonstrated adequate selectivity and stability besides; there was no significant interference observed from other probable protein based components in the real sample matrix. Unlabelled Image • The first label-free electrochemical immunosensor for Candidatus Phytoplasma Aurantifolia (CPA) detection. • Adequate specificity of fabricated immunosensor against interferences from sap samples. • Noticeable LOD that is considerably lower than reported records for CPA detection. [ABSTRACT FROM AUTHOR]