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Microchimica Acta

, Volume 179, Issue 1–2, pp 71–81 | Cite as

Nanoporous gold as a solid support for protein immobilization and development of an electrochemical immunoassay for prostate specific antigen and carcinoembryonic antigen

  • Binod Pandey
  • Alexei V. Demchenko
  • Keith J. StineEmail author
Original Paper

Abstract

Nanoporous gold (NPG) was utilized as a support for immobilizing alkaline phosphatase (ALP) conjugated to monoclonal antibodies against either prostate specific antigen (PSA) or carcinoembryonic antigen (CEA). The antibody-ALP conjugates were coupled to self-assembled monolayers of lipoic acid and used in direct kinetic assays. Using the enzyme substrate p-aminophenylphosphate, the product p-aminophenol was detected by its oxidation near 0.1 V (vs. Ag|AgCl) using square wave voltammetry. The difference in peak current arising from oxidation of p-aminophenol before and after incubation with biomarker increased with biomarker concentration. The response to these two biomarkers was linear up to 10 ng mL−1 for CEA and up to 30 ng mL−1 for PSA. The effect of interference on the PSA assay was studied using bovine serum albumin (BSA) as a model albumin protein. The effect of interference from a serum matrix was examined for the PSA assay using newborn calf serum. A competitive version of the immunoassay using antigen immobilized onto the NPG surface was highly sensitive at lower antigen concentration. Estimates of the surface coverage of the antibody-ALP conjugates on the NPG surface are presented.

Figure

Use of nanoporous gold as a support for a direct kinetic assay of antibody-antigen binding is demonstrated using square-wave voltammetry.

Keywords

Nanoporous gold Immunoassay Self-assembled monolayer Square-wave voltammetry Carcinoembryonic antigen Prostate specific antigen 

Notes

Acknowledgments

The authors thank Professor Fraundorf, Jay K. Bhattarai, Dr. David Osborn and Dr. Dan Zhou of the UM-St. Louis Center for Nanoscience for usage and discussion of SEM. This work was supported by UM-St. Louis and by the NIGMS award R01-GM090254.

Supplementary material

604_2012_870_MOESM1_ESM.docx (159 kb)
ESM 1 (DOCX 158 kb)

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Copyright information

© Springer-Verlag 2012

Authors and Affiliations

  • Binod Pandey
    • 1
  • Alexei V. Demchenko
    • 2
  • Keith J. Stine
    • 1
    Email author
  1. 1.Department of Chemistry and Biochemistry, Center for NanoscienceUniversity of Missouri- Saint LouisSt. LouisUSA
  2. 2.Department of Chemistry and BiochemistryUniversity of Missouri- Saint LouisSt. LouisUSA

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