An economic, square-shaped flat-field illumination module for TIRF-based super-resolution microscopy

Jeff Y L Lam; Yunzhao Wu; Eleni Dimou; Ziwei Zhang; Matthew R Cheetham; Markus Körbel; Zengjie Xia; David Klenerman; John S H Danial

doi:10.1016/j.bpr.2022.100044

An economic, square-shaped flat-field illumination module for TIRF-based super-resolution microscopy

Biophys Rep (N Y). 2022 Mar 9;2(1):None. doi: 10.1016/j.bpr.2022.100044.

Authors

Jeff Y L Lam^{1

2}, Yunzhao Wu^{1

2}, Eleni Dimou^{1

2}, Ziwei Zhang¹, Matthew R Cheetham^{1

2}, Markus Körbel¹, Zengjie Xia^{1

2}, David Klenerman^{1

2}, John S H Danial^{1

2}

Affiliations

¹ Yusuf Hamied Department of Chemistry, University of Cambridge, Cambridge CB2 1EW, United Kingdom.
² UK Dementia Research Institute, University of Cambridge, Cambridge CB2 0AH, United Kingdom.

Abstract

Super-resolution microscopy allows complex biological assemblies to be observed with remarkable resolution. However, the presence of uneven Gaussian-shaped illumination hinders its use in quantitative imaging or high-throughput assays. Methods developed to circumvent this problem are often expensive, hard to implement, or not applicable to total internal reflection fluorescence imaging. We herein demonstrate a cost-effective method to overcome these challenges using a small square-core multimodal optical fiber as the coupler. We characterize our method with synthetic, recombinant, and cellular systems imaged under total internal reflection fluorescence and highly inclined and laminated optical sheet illuminations to demonstrate its ability to produce highly uniform images under all conditions.