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Research Article

Ratiometric fluorescent Si-FITC nanoprobe for immunoassay of SARS-CoV-2 nucleocapsid protein

Guobin Mao1,§Silu Ye1,2,§Wen Yin2Yang Yang3Xinghu Ji4Jin He2Yingxia Liu3Junbiao Dai1Zhike He4( )Yingxin Ma1( )
CAS Key Laboratory of Quantitative Engineering Biology, Guangdong Provincial Key Laboratory of Synthetic Genomics and Shenzhen Key Laboratory of Synthetic Genomics, Shenzhen Institute of Synthetic Biology, Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences, Shenzhen 518055, China
State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan 430070, China
Shenzhen Key Laboratory of Pathogen and Immunity, National Clinical Research Center for Infectious Disease, State Key Discipline of Infectious Disease, Shenzhen Third People's Hospital, Second Hospital Affiliated to Southern University of Science and Technology, Shenzhen 518112, China
College of Chemistry and Molecular Sciences, Wuhan University, Wuhan 430072, China

§ Guobin Mao and Silu Ye contributed equally to this work.

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Graphical Abstract

A ratiometric fluorescent probe, Si-FITC NPs, was prepared via a one-pot hydrothermal method for immunoassay of SARS-CoV-2 N protein. The sensitivity of the ratiometric ELISA was higher than that of the commercial kit and the COVID-19 and non-COVID-19 samples were effectively distinguished by the ratiometric ELISA.


Coronavirus disease 2019 (COVID-19) highlights the importance of rapid and reliable diagnostic assays for the management of virus transmission. Here, we developed a one-pot hydrothermal method to prepare Si-FITC nanoparticles (NPs) for the fluorescent immunoassay of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid protein (N protein). The synthesis of Si-FITC NPs did not need post-modification, which addressed the issue of quantum yield reduction during the coupling reaction. Si-FITC NPs showed two distinct peaks, Si fluorescence at λem = 385 nm and FITC fluorescence at λem = 490 nm. In the presence of KMnO4, Si fluorescence was decreased and FITC fluorescence was enhanced. Briefly, in the presence of N protein, catalase (CAT)-linked secondary antibody/reporter antibody/N protein/capture antibody immunocomplexes were formed on microplates. Subsequently, hydrogen peroxide (H2O2) and Si-FITC NPs/KMnO4 were injected into the microplate together. The decomposition of H2O2 by CAT resulted in remaining of KMnO4, which changed the fluorescence intensity ratio of Si-FITC NPs. The fluorescence intensity ratio correlated significantly with the N protein concentration ranging from 0.02 to 50.00 ng/mL, and the detection limit was 0.003 ng/mL, which was more sensitive than the commercial ELISA kit with a detection limit of 0.057 ng/mL. The N protein concentration can be accurately determined in human serum. Furthermore, the COVID-19 and non-COVID-19 patients were distinguishable by this method. Therefore, the ratiometric fluorescent immunoassay can be used for SARS-CoV-2 infection diagnosis with a high sensitivity and selectivity.

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Nano Research
Pages 2859-2865
Cite this article:
Mao G, Ye S, Yin W, et al. Ratiometric fluorescent Si-FITC nanoprobe for immunoassay of SARS-CoV-2 nucleocapsid protein. Nano Research, 2023, 16(2): 2859-2865.






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Received: 04 July 2022
Revised: 31 August 2022
Accepted: 01 September 2022
Published: 30 September 2022
© Tsinghua University Press 2022