1. State Key Laboratory of Food Science and Technology, School of Food Science and Technology, Nanchang University, Nanchang 330047, P. R. China
2. Beijing Engineering and Technology Research Center of Food Additives, Beijing Technology & Business University, Beijing 100048, P. R. China
3. Departments of Diagnostic Radiology, Surgery, Chemical and Biomolecular Engineering, and Biomedical Engineering, Yong Loo Lin School of Medicine and Faculty of Engineering, National University of Singapore, Singapore, 119074, Singapore
4. Clinical Imaging Research Centre, Centre for Translational Medicine, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117599, Singapore
5. Nanomedicine Translational Research Program, NUS Center for Nanomedicine, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117597, Singapore
*These authors contributed equally to this work.
Background: Gold nanoparticle-based immunochromatographic assay (AuNP-ICA) has insufficient sensitivity due to its inherent colorimetric signal intensity and low capture efficiency of AuNPs. The metal in situ growth is a common strategy to enhance the sensitivity of AuNP-ICA due to its superior signal amplification potential and simple operation. However, the detection distortion caused by metal self-nucleation during the growth process can seriously affect the accuracy and reproducibility of the strips.
Methods: We present a pH-regulated gold in situ growth (GISG) strategy to amplify the colorimetric signal and demonstrate its application in improving the performance of traditional AuNP-ICA. The controllable growth signal amplification is achieved by lowering the pH of the growth solution to weaken the reducibility of hydroxylamine (HA), thus urging the crystallization and growth of Au3+ on the AuNP surface instead of free reduction and self-nucleation. In addition, the mechanism of pH regulation on HA reducibility is elucidated by introducing an electron-donating or electron-withdrawing group to affect the electron density of hydroxyl group.
Results: The proposed GISG strategy shows improved sensitivity, low background, robust operation, and good reproducibility. The LOD values of the designed GISG-amplified AuNP-ICA are as low as 0.0198 ng mL-1 for hepatitis B surface antigen and 0.0125 ng mL-1 for HIV-1 capsid p24 antigen, which are lower by about 500- and 70-fold, respectively, than those of the unamplified AuNP-ICA.
Conclusions: This method is extended to enable ultrasensitive and rapid diagnosis of viral infections, and has potential as a general signal amplification platform to redefine immunochromatographic diagnostics.
Keywords: Immunochromatographic assay, Gold in situ growth, Hydroxylamine, Signal amplification