JoVE Logo
Faculty Resource Center

Sign In





Representative Results






Real-Time Detection of Dynamic Affinity between Biomolecules Using Surface Plasmon Resonance (SPR) Technology

Published: September 29th, 2023



1School of Basic Medical Sciences, Chengdu University of Traditional Chinese Medicine, 2Innovative Institute of Chinese Medicine and Pharmacy, Chengdu University of Traditional Chinese Medicine, 3Research Institute of Integrated Traditional Chinese Medicine and Western Medicine, Chengdu University of Traditional Chinese Medicine
* These authors contributed equally

The present study aims to elucidate the principle and methodology of surface plasmon resonance (SPR) technology, which finds versatile applications across multiple domains. This article describes SPR technology, its operational simplicity, and its remarkable efficacy, with the goal of fostering broader awareness and adoption of this technology among readers.

Surface plasmon resonance (SPR) technology is a sensitive precise method for detecting viruses, pathogenic molecular proteins, and receptors, determining blood types, and detecting food adulteration, among other biomolecular detections. This technology allows for the rapid identification of potential binding between biomolecules, facilitating fast and user-friendly, non-invasive screening of various indicators without the need for labeling. Additionally, SPR technology facilitates real-time detection for high-throughput drug screening. In this program, the application field and basic principles of SPR technology are briefly introduced. The operation process is outlined in detail, starting with instrument calibration and basic system operation, followed by ligand capture and multi-cycle analysis of the analyte. The real-time curve and experimental results of binding quercetin and calycosin to KCNJ2 protein were elaborated upon. Overall, SPR technology provides a highly specific, simple, sensitive, and rapid method for drug screening, real-time detection of related pharmacokinetics, virus detection, and environmental and food safety identification.

Surface plasmon resonance (SPR) technology is an optical detection technique that eliminates the need for labeling the analyte. It enables real-time and dynamic monitoring of quantitative binding affinity, kinetics, and thermodynamics. This high-throughput capacity is highly sensitive and reproducible, allowing for the measurement of various open rates, off rates, and affinity. Additionally, the small sample quantity required further enhances the utility of this method1,2. The fast response biomolecular detection method3, which monitors the affinity binding between....

Log in or to access full content. Learn more about your institution’s access to JoVE content here

NOTE: The complete experimental sensing curve indicates that the experimental process can be categorized into eight distinct stages.

1. Sample and buffer preparation

  1. Prepare sensor chips before the experiment.
    1. Treat the chips with the piranha solution (30% H2O2: H2SO4=1:3; v/v) for 2 min. Subsequently, clean the chip thoroughly with a large amount of deionized water and then soak it in anhydrous ethanol, allow.......

Log in or to access full content. Learn more about your institution’s access to JoVE content here

To determine whether the protein is fixed on the chip surface, the ordinate (response signal) of the SPR sensor map (Figure 1) is used, while the angular displacement of the SPR curve is obtained. Figure 2 and Figure 3 depict the SPR curve of the interaction between quercetin and calycosin with KCNJ2 recombinant protein on the immobilized surface of KCNJ2 recombinant protein after control reduction at concentrations ranging from 3.9.......

Log in or to access full content. Learn more about your institution’s access to JoVE content here

The SPR analysis cycle is divided into four stages. The first stage, the baseline, involves the injection of the buffer. Following that is the second stage, ligand capturing. The sensor chip COOH is activated with EDC/NHS (1:1) at a flow rate of 20 µL/min. The chip is then deactivated using 1 M ethanolamine hydrochloride-NaOH at a flow rate of 20 µL/min. Moving on to the third stage, the multi-cycle analyte method. The analyte is injected into the channel at a flow rate of 20 µL/min for an association phas.......

Log in or to access full content. Learn more about your institution’s access to JoVE content here

This work was supported by the Sichuan Provincial Major R&D Project (2022YFS043), the Key Research and Development Program of Ningxia (2023BEG02012), and Xinglin Scholar Research Promotion Project of Chengdu University of TCM (XKTD2022013).


Log in or to access full content. Learn more about your institution’s access to JoVE content here

NameCompanyCatalog NumberComments
1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC)Nan Jing Reagent,Nanjing,ChinaC08296594
Anhydrous ethanolMerck Chemical Technologies Ltd., Shanghai, China459836
BIAnormalizing solutionMerck Chemical Technologies Ltd., Shanghai, China49781
Blocking solutionBosheng Biotechnology Co.,Ltd., Shanghai, China110050
Bromoacetic acidMerck Chemical Technologies Ltd., Shanghai, China17000
CalycosinPush Bio-technology Co., Ltd., Chengdu, ChinaPU0124-0025
DextranCanspec Scientific Instruments Co., Ltd.,Shanghai, ChinaPM10036
EpichlorohydrinMerck Chemical Technologies Ltd., Shanghai, China492515
Ethanolamine hydrochlorideYuanye Biotech Co., Ltd., Shanghai, ChinaS44235
Glycine-HClMerck Chemical Technologies Ltd., Shanghai, ChinaG2879
H2O2Merck Chemical Technologies Ltd., Shanghai, China3587191
H2SO4Nantong high-tech Industrial Development Zone,China2020001150C
HEPESXiya Reagent Co., Ltd., Shandong, ChinaS3872
KCNJ2 (Human) Recombinant ProteinAbnova,West Meijie Technology Co., Ltd., Beijing, ChinaH00003759-Q01
MUOHJizhi Biochemical Technology Co., Ltd., Shanghai, ChinaM40590
NaOHMerck Chemical Technologies Ltd., Shanghai, ChinaSX0603
N-Hydroxysuccinimide(NHS)Yuanye Biotech Co., Ltd., Shanghai, ChinaS13005
QuercetinPush Bio-technology Co., Ltd., Chengdu, ChinaPU0041-0025
Sensor Chip COOHNicoya
Sodium AcetateMerck Chemical Technologies Ltd., Shanghai, China229873

  1. Jebelli, A., Oroojalian, F., Fathi, F., Mokhtarzadeh, A., Guardia, M. Recent advances in surface plasmon resonance biosensors for microRNAs detection. Biosens Bioelectron. 169, 112599 (2020).
  2. Sun, B., Xu, J., Liu, S., Li, Q. X. Characterization of small molecule-protein interactions using SPR method. Methods Mol Biol. 2690, 149-159 (2023).
  3. Mousavi, S. M., et al. Biomedical applications of an ultra-sensitive surface plasmon resonance biosensor based on smart MXene quantum dots (SMQDs). Biosensors (Basel). 12 (9), 743 (2022).
  4. Olaru, A., Bala, C., Jaffrezic-Renault, N., Aboul-Enein, H. Y. Surface plasmon resonance (SPR) biosensors in pharmaceutical analysis. Crit Rev Anal Chem. 45 (2), 97-105 (2015).
  5. Meschendoerfer, W., Gassner, C., Lipsmeier, F., Regula, J. T., Moelleken, J. SPR-based assays enable the full functional analysis of bispecific molecules. J Pharm Biomed Anal. 132, 141-147 (2017).
  6. Djaileb, A., et al. Cross-validation of ELISA and a portable surface plasmon resonance instrument for IgG antibody serology with SARS-CoV-2 positive individuals. Analyst. 146 (15), 4905-4917 (2021).
  7. Miyake, S., et al. Simultaneous detection of six different types of pesticides by an immunosensor based on surface plasmon resonance. Anal Sci. 36 (3), 335-340 (2020).
  8. Ravindran, N., et al. Recent advances in surface plasmon resonance (SPR) biosensors for food analysis: a review. Crit Rev Food Sci Nutr. 63 (8), 1055-1077 (2023).
  9. Bhandari, D., Chen, F. C., Bridgman, R. C. Magnetic nanoparticles enhanced surface plasmon resonance biosensor for rapid detection of Salmonella typhimurium in Romaine lettuce. Sensors (Basel). 22 (2), 475 (2022).
  10. Amirjani, A., Kamani, P., Hosseini, H. R. M., Sadrnezhaad, S. K. SPR-based assay kit for rapid determination of Pb2. Anal Chim Acta. 1220, 340030 (2022).
  11. Shrivastav, A. M., Cvelbar, U., Abdulhalim, I. A comprehensive review on plasmonic-based biosensors used in viral diagnostics. Commun Biol. 4 (1), 70 (2021).
  12. Nguyen, V. T., et al. Highly sensitive sandwich-type SPR based detection of whole H5Nx viruses using a pair of aptamers. Biosens Bioelectron. 86, 293-300 (2016).
  13. Chang, Y. F., et al. Simple strategy for rapid and sensitive detection of avian influenza A H7N9 virus based on intensity-modulated SPR biosensor and new generated antibody. Anal Chem. 90 (3), 1861-1869 (2018).
  14. Das, C. M., Guo, Y., Kang, L., Ho, H. P., Yong, K. T. Investigation of plasmonic detection of human respiratory virus. Adv Theory Simul. 3 (7), 2000074 (2020).
  15. Lakayan, D., Haselberg, R., Niessen, W. M., Somsen, G. W., Kool, J. On-line coupling of surface plasmon resonance optical sensing to size-exclusion chromatography for affinity assessment of antibody samples. J Chromatogr. A. 1452, 81-88 (2016).
  16. Loo, J. F., et al. An aptamer bio-barcode (ABC) assay using SPR, RNase H, and probes with RNA and gold-nanorods for anti-cancer drug screening. Analyst. 142 (19), 3579-3587 (2017).
  17. Fabini, E., Danielson, U. H. Monitoring drug-serum protein interactions for early ADME prediction through surface plasmon resonance technology. J Pharm Biomed Anal. 144, 188-194 (2017).
  18. Khalenkov, A. M., Norton, M. G., Scoot, D. E. Method for screening influenza neutralizing antibodies in crude human plasma and its derivatives using SPR. Heliyon. 9 (5), 15651 (2023).
  19. Locatelli-Hoops, S., Yeliseev, A. A., Gawrisch, K., Gorshkova, I. Surface plasmon resonance applied to G protein-coupled receptors. Biomed Spectrosc Imaging. 2 (3), 155-181 (2013).
  20. Singh, S., et al. 2D nanomaterial-based surface plasmon resonance sensors for biosensing applications. Micromachines (Basel). 11 (8), 779 (2020).
  21. Pourmadadi, M., et al. Properties and applications of graphene and its derivatives in biosensors for cancer detection: A comprehensive review. Biosensors. 12 (5), 269 (2022).
  22. Singh, T. I., Singh, P., Karki, B. Early detection of chikungunya virus utilizing the surface plasmon resonance comprising a silver-silicon-PtSe 2 multilayer structure. Plasmonics. 18 (3), 1173-1180 (2023).
  23. Das, S., Devireddy, R., Gartia, M. R. Surface plasmon resonance (SPR) sensor for cancer biomarker detection. Biosensors (Basel. 13 (3), 396 (2023).
  24. Liu, R., Ye, X., Cui, T. Recent progress of biomarker detection sensors. Research (Wash D C). 2020, 7949037 (2020).
  25. Bolognesi, M., et al. A fully integrated miniaturized optical biosensor for fast and multiplexing plasmonic detection of high- and low-molecular-weight analytes. Adv Mater. 35 (26), 2208719 (2023).
  26. Inoue, S., Fukada, K., Hayashi, K., Seyama, M. Data processing of SPR curve data to maximize the extraction of changes in electrochemical SPR measurements. Biosensors (Basel). 12 (8), 615 (2022).
  27. Topor, C. V., Puiu, M., Bala, C. Strategies for surface design in surface plasmon resonance (SPR) sensing. Biosensors (Basel). 13 (4), 465 (2023).
  28. Bonnet, H., et al. Negative SPR signals during low molecular weight analyte recognition). Anal Chem. 93 (8), 4134-4140 (2021).
  29. He, P., et al. Cholesterol chip for the study of cholesterol-protein interactions using SPR. Biosensors (Basel). 12 (10), 788 (2022).
  30. Kausaite-Minkstimiene, A., et al. An ultra-sensitive SPR immunosensor for quantitative determination of human cartilage oligomeric matrix protein biomarker. Biosens Bioelectron. 234, 115370 (2023).
  31. Pandey, P. S., et al. SPR based biosensing chip for COVID-19 diagnosis-A review. IEEE Sens J. 22 (14), 13800-13810 (2022).
  32. Mei, Y., et al. Single-layer graphene-coated gold chip for electrochemical surface plasmon resonance study. Anal Bioanal Chem. 411, 4577-4585 (2019).
  33. Zhou, L., Arugula, M. A., Chin, B. A., Simonian, A. L. Simultaneous surface plasmon resonance/fluorescence spectroelectrochemical in situ monitoring of dynamic changes on functional interfaces: A study of the electrochemical proximity assay model system. ACS Appl Mater Interfaces. 10 (48), 41763-41772 (2018).


Surface Plasmon Resonance

This article has been published

Video Coming Soon

JoVE Logo


Terms of Use





Copyright © 2024 MyJoVE Corporation. All rights reserved