A Highly-Sensitive Colorimetric Assay Method for Antibody Array Based on an Tyramide Signal Amplification System

Abstract

In this work, a highly-sensitive and cost-effective detection approach based on the integration of tyramide signal amplification with a silver enhancement method (SEM-TSA) has been developed successfully. To demonstrate the feasibility of this approach, human IgG as a model target protein was employed and its concentration was assayed based on colorimetric detection. The analytical parameters including the concentrations of detection antibody, streptavidin-horseradish peroxidase, biotinyl tyramide, and streptavidin-nanogold were systematically optimized. The quantitative analysis was performed and a dynamic range was obtained from 0.18 ng/mL to 39.1 ng/mL, while no detectable images could be observed when the silver enhancement method (SEM) without TSA was used. The detection limits were 0.18 ng/mL and 21 ng/mL for SEM-TSA and SEM, respectively. The results showed that sensitivity of the presented colorimetric assay significantly increased by two-orders of magnitude. In addition, this method has been successfully applied in analyzing normal human serum samples. The results suggested the colorimetric detection method based on TSA-SEM has promising potential applications in biomedical analysis and clinical diagnosis.

Keywords:

• Antibody arrays
• Colorimetric assay
• Silver enhancement method
• Tyramide signal amplification

Acknowledgments

The authors acknowledge financial support of National Natural Foundation of China (Grant No. 20975050, 21175066), the National High-Tech Development Plan (863 Program, 2009AA093701), Jiangsu Province Science and Technology Support Program (No. BE2011773), Research Foundation of Jiangsu Province Environmental Monitoring (No. 1116), National Basic Research Program of China (973 Program, No. 2011CB911003), and the National Science Funds for Creative Research Groups (No. 21121091).

Notes

*Analytical conditions are as described in experimental section.

This paper was submitted as part of a Special Issue on Biosensors organized by Dr. Yu Lei of the University of Connecticut.

Xiaobo Yu is currently at the Virginia G. Piper Center for Personalized Diagnostics, Biodesign Institute, Arizona State University, Tempe Arizona, USA. Maika Vuki is currently at the college of Natural and Applied Sciences, University of Guam, Mangilao, Guam, USA

Additional information

Notes on contributors

Xiaobo Yu

Xiaobo Yu is currently at the Virginia G. Piper Center for Personalized Diagnostics, Biodesign Institute, Arizona State University, Tempe, Arizona, USA.

Maika Vuki

Maika Vuki is currently at the College of Natural and Applied Sciences, University of Guam, Mangilao, Guam, USA