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Nano Research 2022, 15(4): 3487-3495 https://doi.org/10.1007/s12274-021-3902-1
Research Article | Issue | Published: 20 October 2021

Multiplex ratiometric gold nanoprobes based on surface-enhanced Raman scattering enable accurate molecular detection and imaging of bladder cancer

Show Author's Information Xiao Liang1,2,§,  Pu Zhang1,3,§,  Minghai Ma1,§,  Tao Yang1,  Xiangwei Zhao4,  Rui Zhang1,  Minxuan Jing1,  Rundong Song1,  Lei Wang2( ),  Jinhai Fan1( )
Department of Urology, The First Affiliated Hospital, Xi’an Jiaotong University, Xi’an 710061, China
Department of Thoracic Surgery, Tangdu Hospital, Air Force Medical University, Xi’an 710038, China
Department of Urology, Zhejiang Provincial People’s Hospital, People’s Hospital of Hangzhou Medical College, Hangzhou 310014, China
State Key Laboratory of Bioelectronics, School of Biological Science and Medical Engineering, National Demonstration Center for Experimental Biomedical Engineering Education, Southeast University, Nanjing 210096, China

§ Xiao Liang, Pu Zhang, and Minghai Ma contributed equally to this work.

Keywords:
surface-enhanced Raman scattering (SERS), internal reference, bladder cancer, non-invasive detection, SERS imaging
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Nano detection
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Liang X, Zhang P, Ma M, et al. Multiplex ratiometric gold nanoprobes based on surface-enhanced Raman scattering enable accurate molecular detection and imaging of bladder cancer. Nano Research, 2022, 15(4): 3487-3495. https://doi.org/10.1007/s12274-021-3902-1
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Abstract Full text Electronic supplementary material About this article

Recently, surface-enhanced Raman scattering (SERS) has been successfully used in the non-invasive detection of bladder tumor (BCa). The internal standard method was considered as an effective ratiometric strategy for calibrating signal fluctuation originated from the interference of measurement conditions and samples. However, it is still difficult to detect the target mRNA quantitatively using the current ratiometric SERS nanosensors. In this study, we developed an internal reference based ratiometric SERS assay. Two kinds of molecular beacons (MB) carrying Raman reporter molecules were anchored on sea-urchin-like Au nanoclusters (AuNCs). Thymidine kinase1 (TK1) MBs with hexachlorofluorescein (HEX) were used to capture tumor biomarker TK1 mRNA, and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) MBs with 5(6)-carboxyfluorescein (FAM) were used to offer internal standard signals. The internal reference GAPDH MB can reflect the consistent content of the GAPDH mRNA in single cells. The ratiometric method (I745/I645) can more accurately reflect the content of target mRNA in single cells. The ratiometric nanoprobes had excellent stability (coefficient of variation: 0.3%), high sensitivity (detection limit: 3.4 pM), high specificity (capable of single-base mismatch recognition) and ribozyme-resistant stability. Notably, the nanoprobes can effectively distinguish BCa cells from normal cells, and it was easy to contour the single BCa cell using the ratiometric method. By combining asymmetric polymerase chain reaction (PCR) and ratiometric nanoprobes, it was easy to distinguish the SERS ratio (I745/I645) as low concentration as 10−14 M. Further clinical detection in urine samples from patients with BCa confirmed its potential for early noninvasive diagnosis of BCa with the sensitivity of 80% and specificity of 100%, which is superior to the current urine cytological method.


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Abstract
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Electronic supplementary material
About this article

Multiplex ratiometric gold nanoprobes based on surface-enhanced Raman scattering enable accurate molecular detection and imaging of bladder cancer

Show Author's information Xiao Liang1,2,§, Pu Zhang1,3,§, Minghai Ma1,§, Tao Yang1, Xiangwei Zhao4, Rui Zhang1, Minxuan Jing1, Rundong Song1, Lei Wang2( ), Jinhai Fan1( )
Department of Urology, The First Affiliated Hospital, Xi’an Jiaotong University, Xi’an 710061, China
Department of Thoracic Surgery, Tangdu Hospital, Air Force Medical University, Xi’an 710038, China
Department of Urology, Zhejiang Provincial People’s Hospital, People’s Hospital of Hangzhou Medical College, Hangzhou 310014, China
State Key Laboratory of Bioelectronics, School of Biological Science and Medical Engineering, National Demonstration Center for Experimental Biomedical Engineering Education, Southeast University, Nanjing 210096, China

§ Xiao Liang, Pu Zhang, and Minghai Ma contributed equally to this work.

Abstract

Recently, surface-enhanced Raman scattering (SERS) has been successfully used in the non-invasive detection of bladder tumor (BCa). The internal standard method was considered as an effective ratiometric strategy for calibrating signal fluctuation originated from the interference of measurement conditions and samples. However, it is still difficult to detect the target mRNA quantitatively using the current ratiometric SERS nanosensors. In this study, we developed an internal reference based ratiometric SERS assay. Two kinds of molecular beacons (MB) carrying Raman reporter molecules were anchored on sea-urchin-like Au nanoclusters (AuNCs). Thymidine kinase1 (TK1) MBs with hexachlorofluorescein (HEX) were used to capture tumor biomarker TK1 mRNA, and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) MBs with 5(6)-carboxyfluorescein (FAM) were used to offer internal standard signals. The internal reference GAPDH MB can reflect the consistent content of the GAPDH mRNA in single cells. The ratiometric method (I745/I645) can more accurately reflect the content of target mRNA in single cells. The ratiometric nanoprobes had excellent stability (coefficient of variation: 0.3%), high sensitivity (detection limit: 3.4 pM), high specificity (capable of single-base mismatch recognition) and ribozyme-resistant stability. Notably, the nanoprobes can effectively distinguish BCa cells from normal cells, and it was easy to contour the single BCa cell using the ratiometric method. By combining asymmetric polymerase chain reaction (PCR) and ratiometric nanoprobes, it was easy to distinguish the SERS ratio (I745/I645) as low concentration as 10−14 M. Further clinical detection in urine samples from patients with BCa confirmed its potential for early noninvasive diagnosis of BCa with the sensitivity of 80% and specificity of 100%, which is superior to the current urine cytological method.

Keywords: surface-enhanced Raman scattering (SERS), internal reference, bladder cancer, non-invasive detection, SERS imaging

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Publication history
Copyright
Acknowledgements

Publication history

Received: 22 August 2021
Revised: 19 September 2021
Accepted: 22 September 2021
Published: 20 October 2021
Issue date: April 2022

Copyright

© Tsinghua University Press and Springer-Verlag GmbH Germany, part of Springer Nature 2021

Acknowledgements

Acknowledgements

This work was supported by the National Natural Science Foundation of China (No. 81901838), the Key Research and Development Plan in Shaanxi Province (Nos. 2020SF-123 and 2020SF-195), the Natural Science Foundation of Zhejiang Province (No. LQ21H160041), and the Medical Research Program of Department of Science and Technology of Xi’an, Shaanxi Province (No. 2019115713YX012SF048(4)). We thank Mengzhao Zhang, Qiuya Shao, Lu Wang and Lu Zhang at Department of Urology, the First Affiliated Hospital, Xi’an Jiaotong University for their support. We also thank Dr. Yu Wang at Instrument Analysis Center of Xi’an Jiaotong University for her assistance.

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