Detection of C‐Reactive Protein by Electrochemiluminescence‐Based Droplet‐Free Digital Immunoassay with Tyramide Signal Amplification
C‐reactive protein (CRP) is a widely used biomarker for monitoring inflammatory processes and assessing disease progression in various conditions, including cardiovascular diseases, infections, and autoimmune disorders. In this study, a simplified droplet‐free digital enzyme‐linked immunosorbent ass...
Saved in:
| Main Authors: | , , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Wiley-VCH
2025-06-01
|
| Series: | ChemElectroChem |
| Subjects: | |
| Online Access: | https://doi.org/10.1002/celc.202500024 |
| Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
| _version_ | 1850160324927291392 |
|---|---|
| author | Hiroya Abe Shuri Nakamura Kentaro Ito Kosuke Ino Zhengyi Liu Shinichi Sato Kumi Y. Inoue Hitoshi Shiku |
| author_facet | Hiroya Abe Shuri Nakamura Kentaro Ito Kosuke Ino Zhengyi Liu Shinichi Sato Kumi Y. Inoue Hitoshi Shiku |
| author_sort | Hiroya Abe |
| collection | DOAJ |
| description | C‐reactive protein (CRP) is a widely used biomarker for monitoring inflammatory processes and assessing disease progression in various conditions, including cardiovascular diseases, infections, and autoimmune disorders. In this study, a simplified droplet‐free digital enzyme‐linked immunosorbent assay (ELISA) system is developed based on electrochemiluminescence (ECL) imaging for the sensitive detection of CRP. This system eliminates the need for droplet or microwell arrays, significantly simplifying the workflow while maintaining high sensitivity. Tyramide signal amplification was employed to enhance signal generation, relying on its localized reaction to achieve precise single‐particle amplification with minimal signal diffusion. The developed assay is capable of detecting poly‐horseradish peroidase at concentrations as low as 1.0 fmol L−1 and successfully adhered to the Poisson distribution model for digital quantification. The application of this system to CRP detection yields a limit of detection of 17.0 pg mL−1 (162 fmol L−1). The ECL bead ratio increases proportionally with CRP concentration, enabling accurate quantification. Moreover, the system exhibits high specificity, showing no response to other biomolecules and maintaining precise detection even in complex biological samples such as human serum. This study highlights the potential of the droplet‐free digital ELISA system for clinical diagnostics and the detection of low‐abundance biomarkers. |
| format | Article |
| id | doaj-art-0e8cdba1e77e496fb1bfedc67053e701 |
| institution | OA Journals |
| issn | 2196-0216 |
| language | English |
| publishDate | 2025-06-01 |
| publisher | Wiley-VCH |
| record_format | Article |
| series | ChemElectroChem |
| spelling | doaj-art-0e8cdba1e77e496fb1bfedc67053e7012025-08-20T02:23:11ZengWiley-VCHChemElectroChem2196-02162025-06-011212n/an/a10.1002/celc.202500024Detection of C‐Reactive Protein by Electrochemiluminescence‐Based Droplet‐Free Digital Immunoassay with Tyramide Signal AmplificationHiroya Abe0Shuri Nakamura1Kentaro Ito2Kosuke Ino3Zhengyi Liu4Shinichi Sato5Kumi Y. Inoue6Hitoshi Shiku7Graduate School of Engineering Tohoku University Sendai Miyagi 980‐8577 JapanGraduate School of Engineering Tohoku University Sendai Miyagi 980‐8577 JapanGraduate School of Environmental Studies Tohoku University Sendai Miyagi 980‐8577 JapanGraduate School of Engineering Tohoku University Sendai Miyagi 980‐8577 JapanFrontier Research Institute for Interdisciplinary Sciences Tohoku University 6‐3 Aoba, Aramaki, Aoba‐ku Sendai Miyagi 980‐8578 JapanFrontier Research Institute for Interdisciplinary Sciences Tohoku University 6‐3 Aoba, Aramaki, Aoba‐ku Sendai Miyagi 980‐8578 JapanGraduate School of Environmental Studies Tohoku University Sendai Miyagi 980‐8577 JapanGraduate School of Engineering Tohoku University Sendai Miyagi 980‐8577 JapanC‐reactive protein (CRP) is a widely used biomarker for monitoring inflammatory processes and assessing disease progression in various conditions, including cardiovascular diseases, infections, and autoimmune disorders. In this study, a simplified droplet‐free digital enzyme‐linked immunosorbent assay (ELISA) system is developed based on electrochemiluminescence (ECL) imaging for the sensitive detection of CRP. This system eliminates the need for droplet or microwell arrays, significantly simplifying the workflow while maintaining high sensitivity. Tyramide signal amplification was employed to enhance signal generation, relying on its localized reaction to achieve precise single‐particle amplification with minimal signal diffusion. The developed assay is capable of detecting poly‐horseradish peroidase at concentrations as low as 1.0 fmol L−1 and successfully adhered to the Poisson distribution model for digital quantification. The application of this system to CRP detection yields a limit of detection of 17.0 pg mL−1 (162 fmol L−1). The ECL bead ratio increases proportionally with CRP concentration, enabling accurate quantification. Moreover, the system exhibits high specificity, showing no response to other biomolecules and maintaining precise detection even in complex biological samples such as human serum. This study highlights the potential of the droplet‐free digital ELISA system for clinical diagnostics and the detection of low‐abundance biomarkers.https://doi.org/10.1002/celc.202500024digital enzyme‐linked immunosorbent assaydigital immunoassaysdroplet‐free digital immunoassayselectrochemiluminescencetyramide signal amplifications |
| spellingShingle | Hiroya Abe Shuri Nakamura Kentaro Ito Kosuke Ino Zhengyi Liu Shinichi Sato Kumi Y. Inoue Hitoshi Shiku Detection of C‐Reactive Protein by Electrochemiluminescence‐Based Droplet‐Free Digital Immunoassay with Tyramide Signal Amplification ChemElectroChem digital enzyme‐linked immunosorbent assay digital immunoassays droplet‐free digital immunoassays electrochemiluminescence tyramide signal amplifications |
| title | Detection of C‐Reactive Protein by Electrochemiluminescence‐Based Droplet‐Free Digital Immunoassay with Tyramide Signal Amplification |
| title_full | Detection of C‐Reactive Protein by Electrochemiluminescence‐Based Droplet‐Free Digital Immunoassay with Tyramide Signal Amplification |
| title_fullStr | Detection of C‐Reactive Protein by Electrochemiluminescence‐Based Droplet‐Free Digital Immunoassay with Tyramide Signal Amplification |
| title_full_unstemmed | Detection of C‐Reactive Protein by Electrochemiluminescence‐Based Droplet‐Free Digital Immunoassay with Tyramide Signal Amplification |
| title_short | Detection of C‐Reactive Protein by Electrochemiluminescence‐Based Droplet‐Free Digital Immunoassay with Tyramide Signal Amplification |
| title_sort | detection of c reactive protein by electrochemiluminescence based droplet free digital immunoassay with tyramide signal amplification |
| topic | digital enzyme‐linked immunosorbent assay digital immunoassays droplet‐free digital immunoassays electrochemiluminescence tyramide signal amplifications |
| url | https://doi.org/10.1002/celc.202500024 |
| work_keys_str_mv | AT hiroyaabe detectionofcreactiveproteinbyelectrochemiluminescencebaseddropletfreedigitalimmunoassaywithtyramidesignalamplification AT shurinakamura detectionofcreactiveproteinbyelectrochemiluminescencebaseddropletfreedigitalimmunoassaywithtyramidesignalamplification AT kentaroito detectionofcreactiveproteinbyelectrochemiluminescencebaseddropletfreedigitalimmunoassaywithtyramidesignalamplification AT kosukeino detectionofcreactiveproteinbyelectrochemiluminescencebaseddropletfreedigitalimmunoassaywithtyramidesignalamplification AT zhengyiliu detectionofcreactiveproteinbyelectrochemiluminescencebaseddropletfreedigitalimmunoassaywithtyramidesignalamplification AT shinichisato detectionofcreactiveproteinbyelectrochemiluminescencebaseddropletfreedigitalimmunoassaywithtyramidesignalamplification AT kumiyinoue detectionofcreactiveproteinbyelectrochemiluminescencebaseddropletfreedigitalimmunoassaywithtyramidesignalamplification AT hitoshishiku detectionofcreactiveproteinbyelectrochemiluminescencebaseddropletfreedigitalimmunoassaywithtyramidesignalamplification |