TY - JOUR
T1 - Label-Free Imaging of Single Proteins and Binding Kinetics Using Total Internal Reflection-Based Evanescent Scattering Microscopy
AU - Zhang, Pengfei
AU - Wang, Rui
AU - Wan, Zijian
AU - Zhou, Xinyu
AU - Ma, Guangzhong
AU - Kolay, Jayeeta
AU - Jiang, Jiapei
AU - Wang, Shaopeng
N1 - Publisher Copyright:
© 2022 American Chemical Society.
PY - 2022/8/2
Y1 - 2022/8/2
N2 - Single-molecule detection can push beyond ensemble averages and reveal the statistical distributions of molecular properties. Measuring the binding kinetics of single proteins also represents one of the critical and challenging tasks in protein analysis. Here, we report total internal reflection-based evanescent scattering microscopy with label-free single-protein detection capability. Total internal reflection is employed to excite the evanescent field to enhance light-analyte interaction and reduce environmental noise. As a result, the system provides wide-field imaging capability and allows excitation and observation using one objective. In addition, this system quantifies protein binding kinetics by simultaneously counting the binding of individual molecules and recording their binding sites with nanometer precision, providing a digital method to measure binding kinetics with high spatiotemporal resolution. This approach does not employ specially designed microspheres or nanomaterials and may pave a way for label-free single-protein analysis in conventional microscopy.
AB - Single-molecule detection can push beyond ensemble averages and reveal the statistical distributions of molecular properties. Measuring the binding kinetics of single proteins also represents one of the critical and challenging tasks in protein analysis. Here, we report total internal reflection-based evanescent scattering microscopy with label-free single-protein detection capability. Total internal reflection is employed to excite the evanescent field to enhance light-analyte interaction and reduce environmental noise. As a result, the system provides wide-field imaging capability and allows excitation and observation using one objective. In addition, this system quantifies protein binding kinetics by simultaneously counting the binding of individual molecules and recording their binding sites with nanometer precision, providing a digital method to measure binding kinetics with high spatiotemporal resolution. This approach does not employ specially designed microspheres or nanomaterials and may pave a way for label-free single-protein analysis in conventional microscopy.
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U2 - 10.1021/acs.analchem.2c01510
DO - 10.1021/acs.analchem.2c01510
M3 - Article
AN - SCOPUS:85135910258
SN - 0003-2700
VL - 94
SP - 10781
EP - 10787
JO - Analytical Chemistry
JF - Analytical Chemistry
IS - 30
ER -