A standardization implementation, calibration, and analysis protocol for spectroscopic single molecule localization microscopy (sSMLM)

Benjamin Brenner; Wei Hong Yeo; Youngseop Lee; Fengyuanshan Xu; Junghun Kweon; Yang Zhang; Cheng Sun; Hao  F. Zhang

doi:10.1117/12.3001856

13 March 2024 A standardization implementation, calibration, and analysis protocol for spectroscopic single molecule localization microscopy (sSMLM)

Benjamin Brenner, Wei Hong Yeo, Youngseop Lee, Fengyuanshan Xu, Junghun Kweon, Yang Zhang, Cheng Sun, Hao F. Zhang

Author Affiliations +

Proceedings Volume PC12849, Single Molecule Spectroscopy and Superresolution Imaging XVII; PC1284902 (2024) https://doi.org/10.1117/12.3001856
Event: SPIE BiOS, 2024, San Francisco, California, United States

Abstract

Spectroscopic single-molecule localization microscopy (sSMLM) combines super-resolution microscopy and spectroscopy. Its single molecule sensitivity and high spectral precision have made it uniquely valuable for several applications, including multicolor imaging, chemical characterization, polarity sensing, and multiplexed single particle tracking. However, widespread adoption is hampered by a lack of standardization in optical implementation, calibration techniques, and image processing. We demonstrate our lab’s efforts to develop tools that simplify adoption and optimize photon efficiency, including protocols for calibration techniques, a user-friendly imageJ plugin for image processing, and a fabricated monolithic beam splitter and prism designed to fit into a microscope body with minimal optical alignment.

Conference Presentation

Citation Download Citation

Benjamin Brenner, Wei Hong Yeo, Youngseop Lee, Fengyuanshan Xu, Junghun Kweon, Yang Zhang, Cheng Sun, and Hao F. Zhang "A standardization implementation, calibration, and analysis protocol for spectroscopic single molecule localization microscopy (sSMLM)", Proc. SPIE PC12849, Single Molecule Spectroscopy and Superresolution Imaging XVII, PC1284902 (13 March 2024); https://doi.org/10.1117/12.3001856

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