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Proximity hybridization triggered rolling-circle amplification for sensitive electrochemical homogeneous immunoassay
Fenglei Gao1, Fuyi Zhou, Songjie Chen
1School of Chemistry and Materials Science, Jiangsu Normal University, Xuzhou 221116, P. R. China. wangpo@jsnu.edu.cn.
The Analyst
|October 21, 2017
Summary
A novel electrochemical immunoassay enables ultrasensitive detection of carcinoembryonic antigen (CEA) using proximity hybridization and rolling circle amplification. This method offers a sensitive, label-free approach for CEA quantification in complex biological samples.
Area of Science:
- Biomedical Engineering
- Analytical Chemistry
- Biosensors
Background:
- Carcinoembryonic antigen (CEA) is a crucial tumor marker for various cancers.
- Ultrasensitive and specific detection methods are vital for early cancer diagnosis and monitoring.
- Existing immunoassay strategies often face challenges with sensitivity, complexity, or non-specific binding.
Purpose of the Study:
- To develop a novel homogeneous electrochemical immunoassay for ultrasensitive CEA detection.
- To leverage target-induced proximity hybridization and rolling circle amplification (RCA) for signal amplification.
- To achieve immobilization-free detection for simplified assay procedures.
Main Methods:
- Utilized an uncharged peptide nucleic acid (PNA) probe labeled with ferrocene (Fc) on an indium tin oxide (ITO) electrode.
- Employed target-induced proximity hybridization of antibodies and a molecular beacon in homogeneous solution.
- Initiated rolling circle amplification (RCA) using the unfolded molecular beacon as a primer.
- Detected CEA via a signal-off electrochemical approach based on Fc-PNA consumption and electrostatic repulsion.
Main Results:
- Achieved ultrasensitive CEA detection in the range of 1 pg mL-1 to 10 ng mL-1.
- Established a low limit of detection (LOD) of 0.49 pg mL-1 for CEA.
- Demonstrated good selectivity, high sensitivity, acceptable accuracy, and versatility for other analytes.
- Validated the assay's practical applicability by successfully detecting CEA in human serum samples.
Conclusions:
- The developed homogeneous electrochemical immunoassay strategy provides a sensitive and efficient platform for CEA detection.
- The combination of proximity hybridization and RCA offers significant signal amplification for ultrasensitive biomarker quantification.
- The immobilization-free and signal-off mechanism simplifies the assay and enhances its practical utility in clinical diagnostics.
