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<p>This study presents the design, optimization, and characterization of a low-cost, paper-based potentiometric immunosensor for the sensitive detection of <i>Brucella abortus</i>. The sensing platform was constructed using a cellulose paper electrode modified with conductive carbon nanotube (CNT) ink, followed by a stepwise functionalization with Protein A and specific polyclonal anti-<i>Brucella abortus</i> antibodies. Surface assembly and morphological properties were comprehensively characterized using Scanning Electron Microscopy (SEM), Atomic Force Microscopy (AFM), and X-ray Photoelectron Spectroscopy (XPS), which confirmed the successful layer-by-layer deposition. Furthermore, Confocal Laser Scanning Microscopy (CLSM) and Super-Resolution Microscopy by the Single Molecule Localization Microscopy (SMLM) reconstruction were employed to validate antibody immobilization and specific antigen binding at the nanoscale, revealing emitter localization below 100&#xa0;nm. Under optimized conditions, the immunosensor demonstrated a linear potentiometric response with a limit of detection (LOD) of 5 CFU/mL before saturation in the electrode. Selectivity tests showed no significant cross-reactivity against <i>Salmonella typhi</i>, <i>Listeria monocytogenes</i>, or <i>Brucella melitensis</i>. While further validation in complex food matrices is required, these results suggest that the proposed paper-based platform offers a promising, cost-effective, and rapid alternative for the decentralized screening of brucellosis in resource-limited settings.&#xa0;</p> Graphical Abstract <p>Schematic representation of the paper-based potentiometric immunosensor assembly and performance. The workflow illustrates the fabrication of the Paper-Based Electrode (PBE) using carbon nanotube (CNT) ink, followed by stepwise functionalization with Protein A and specific anti-<i>Brucella abortus</i> antibodies. The sensor surface is characterized by using Scanning Electron Microscopy (SEM), Atomic Force Microscopy (AFM), and Super-Resolution Microscopy by Single-Molecule Localization Microscopy (SMLM) to validate the nanoscale assembly. The detection mechanism demonstrates a specific potentiometric response to <i>Brucella abortus</i> with a limit of detection (LOD) of 5 CFU/mL before saturation in the electrode, showing no cross-reactivity with <i>Salmonella typhi</i>, <i>Listeria monocytogenes</i>, or <i>Brucella melitensis</i></p> <p></p>

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Paper-Based Potentiometric Immunosensor for Brucella abortus Detection Using Carbon Nanotubes

  • 期刊论文

  • 发表日期：

    2026年5月2日

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  • Gabriela Damián-Morales,
  • Josué David Hernández-Varela,
  • Martha Moreno-Lafont,
  • Rubén López-Santiago,
  • José Jorge Chanona-Pérez

摘要

This study presents the design, optimization, and characterization of a low-cost, paper-based potentiometric immunosensor for the sensitive detection of Brucella abortus. The sensing platform was constructed using a cellulose paper electrode modified with conductive carbon nanotube (CNT) ink, followed by a stepwise functionalization with Protein A and specific polyclonal anti-Brucella abortus antibodies. Surface assembly and morphological properties were comprehensively characterized using Scanning Electron Microscopy (SEM), Atomic Force Microscopy (AFM), and X-ray Photoelectron Spectroscopy (XPS), which confirmed the successful layer-by-layer deposition. Furthermore, Confocal Laser Scanning Microscopy (CLSM) and Super-Resolution Microscopy by the Single Molecule Localization Microscopy (SMLM) reconstruction were employed to validate antibody immobilization and specific antigen binding at the nanoscale, revealing emitter localization below 100 nm. Under optimized conditions, the immunosensor demonstrated a linear potentiometric response with a limit of detection (LOD) of 5 CFU/mL before saturation in the electrode. Selectivity tests showed no significant cross-reactivity against Salmonella typhi, Listeria monocytogenes, or Brucella melitensis. While further validation in complex food matrices is required, these results suggest that the proposed paper-based platform offers a promising, cost-effective, and rapid alternative for the decentralized screening of brucellosis in resource-limited settings. 

Graphical Abstract

Schematic representation of the paper-based potentiometric immunosensor assembly and performance. The workflow illustrates the fabrication of the Paper-Based Electrode (PBE) using carbon nanotube (CNT) ink, followed by stepwise functionalization with Protein A and specific anti-Brucella abortus antibodies. The sensor surface is characterized by using Scanning Electron Microscopy (SEM), Atomic Force Microscopy (AFM), and Super-Resolution Microscopy by Single-Molecule Localization Microscopy (SMLM) to validate the nanoscale assembly. The detection mechanism demonstrates a specific potentiometric response to Brucella abortus with a limit of detection (LOD) of 5 CFU/mL before saturation in the electrode, showing no cross-reactivity with Salmonella typhi, Listeria monocytogenes, or Brucella melitensis