4.8 Article

Competitive-type displacement reaction for direct potentiometric detection of low-abundance protein

Journal

BIOSENSORS & BIOELECTRONICS
Volume 53, Issue -, Pages 465-471

Publisher

ELSEVIER ADVANCED TECHNOLOGY
DOI: 10.1016/j.bios.2013.10.027

Keywords

Competitive-type displacement reaction; Nanogold-labeled antibody; Prostate-specific antigen; Potentiometric immunoassay; Polyethyleneimine-poly(styrene-co-acrylic acid) microsphere

Funding

  1. National 973 Basic Research Program of China [2010CB732403]
  2. National Natural Science Foundation of China [41176079, 21075019]
  3. Doctoral Program of Higher Education of China [20103514120003]
  4. National Science Foundation of Fujian Province [2011J06003]
  5. China-Russia Bilateral Scientific Cooperation Research Program (NSFC/RFBR) [21211120157]
  6. Program for Changjiang Scholars and Innovative Research Team in University [IRT1116]

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Prostate-specific antigen (PSA), one of the indications of possible prostate malignancy, is used as a biomarker for the diagnosis and prognosis of prostate cancer. Herein, we develop a new homogeneous potentiometric immunoassay for sensitive detection of low-concentration PSA without the need of sample separation and washing step. Two nanostructures including positively charged polyethyleneimine-poly(styrene-co-acrylic acid) (PEI-PSAA) nanospheres and negatively charged gold nanoparticles conjugated with anti-PSA antibody (Ab-AuNP) were first synthesized by using mulsifier-free emulsion copolymerization and wet chemistry method, respectively. Thereafter, the as-prepared PEI-PSAA was used as a pseudo hapten for the construction of immunosensing probe based on an electrostatic interaction between PEI-PSAA and Ab-AuNP. Upon target introduction, the added PSA competed with PEI-PASS for Ab-AuNP based on a specific antigen-antibody interaction, and displaced Ab-AuNP from PEI-PASS. The dissociated PEI-PASS was captured through the negatively charged Nafion- modified electrode, thereby resulting in the change of membrane potential. The fabrication process was characterized by using high-resolution transmission electron microscope (HRTEM), scanning electron microscope with energy-dispersive X-ray spectroscopy (SEM-EDX), surface plasmon resonance (SPR) and dynamic laser scattering (DLS) technique. Under optimal conditions, the output signal was indirectly proportional to the concentration of target PSA in the sample and exhibited a dynamic range from 0.1 to 50 ng/mL with a detection limit (LOD) of 0.04 ng/mL. Intra- and inter-assay coefficients of variation (CVs) were 6.8 and 7.5%, respectively. In addition, the methodology was evaluated for analysis of 12 clinical serum samples and showed good accordance between the results obtained by the developed immunosensing protocol and a commercialized enzyme-linked immunosorbent assay (ELISA) method. (C) 2013 Elsevier B.V. All rights reserved.

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