Abstract
Structured illumination microscopy (SIM) is a method that can double the spatial resolution of wide-field fluorescence microscopy in three dimensions by using spatially structured illumination light. In this chapter, we introduce the basic principles of SIM and describe in detail several different implementations based on either a diffraction grating or liquid crystal spatial light modulators. We also describe nonlinear SIM, a method that in theory can achieve unlimited resolution. In addition, we discuss a number of key points important for high-resolution imaging.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Neil MA, Juskaitis R, Wilson T (1997) Method of obtaining optical sectioning by using structured light in a conventional microscope. Opt Lett 22:1905–1907
Gustafsson MG (2000) Surpassing the lateral resolution limit by a factor of two using structured illumination microscopy. J Microsc 198:82–87
Gustafsson MG, Shao L, Carlton PM et al (2008) Three-dimensional resolution doubling in wide-field fluorescence microscopy by structured illumination. Biophys J 94:4957–4970
Goodman JW (2005) Introduction to Fourier optics. Roberts & Co., Englewood
Kner P, Chhun BB, Griffis ER et al (2009) Super-resolution video microscopy of live cells by structured illumination. Nat Methods 6:339–342
Fiolka R, Shao L, Rego EH et al (2012) Time-lapse two-color 3D imaging of live cells with doubled resolution using structured illumination. Proc Natl Acad Sci U S A 109:5311–5315
Shao L, Kner P, Rego EH et al (2011) Super-resolution 3D microscopy of live whole cells using structured illumination. Nat Methods 8:1044–1046
Gustafsson MG (2005) Nonlinear structured-illumination microscopy: wide-field fluorescence imaging with theoretically unlimited resolution. Proc Natl Acad Sci U S A 102:13081–13086
Rego EH, Shao L, Macklin JJ et al (2012) Nonlinear structured-illumination microscopy with a photoswitchable protein reveals cellular structures at 50-nm resolution. Proc Natl Acad Sci U S A 109:E135–E143
Shao L, Isaac B, Uzawa S et al (2008) I5S: wide-field light microscopy with 100-nm-scale resolution in three dimensions. Biophys J 94:4971–4983
Planchon TA, Gao L, Milkie DE et al (2011) Rapid three-dimensional isotropic imaging of living cells using Bessel beam plane illumination. Nat Methods 8:417–423
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2015 Springer Science+Business Media New York
About this protocol
Cite this protocol
Rego, E.H., Shao, L. (2015). Practical Structured Illumination Microscopy. In: Verveer, P. (eds) Advanced Fluorescence Microscopy. Methods in Molecular Biology, vol 1251. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2080-8_10
Download citation
DOI: https://doi.org/10.1007/978-1-4939-2080-8_10
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-2079-2
Online ISBN: 978-1-4939-2080-8
eBook Packages: Springer Protocols