High speed multi-plane super-resolution structured illumination microscopy of living cells using an image-splitting prism

Super-resolution structured illumination microscopy (SR-SIM) can be conducted at video-rate acquisition speeds when combined with high-speed spatial light modulators and sCMOS cameras, rendering it particularly suitable for live cell imaging. If, however, three-dimensional (3D) information is desired, the sequential acquisition of vertical image stacks employed by current setups significantly slows down the acquisition process. In this work we present a multi-plane approach to SR-SIM that overcomes this slowdown via the simultaneous acquisition of multiple object planes, employing a recently introduced multi-plane image splitting prism combined with high-speed SR-SIM illumination. This strategy requires only the introduction of a single optical element and the addition of a second camera to acquire a laterally super-resolved three-dimensional image stack. We demonstrate the performance of multi-plane SR-SIM by applying this instrument to the dynamics of live mitochondrial network.

[1]  Stefan Jakobs,et al.  Bax assembles into large ring‐like structures remodeling the mitochondrial outer membrane in apoptosis , 2016, The EMBO journal.

[2]  Kevin O'Holleran,et al.  Polarization effects on contrast in structured illumination microscopy. , 2012, Optics letters.

[3]  Rainer Heintzmann,et al.  Super-Resolution Structured Illumination Microscopy. , 2017, Chemical reviews.

[4]  Talley J. Lambert,et al.  Multifocus structured illumination microscopy for fast volumetric super-resolution imaging. , 2017, Biomedical optics express.

[5]  J. Mixter.  Fast , 2012 .

[6]  Clemens F Kaminski,et al.  Speed limits of structured illumination microscopy. , 2017, Optics letters.

[7]  Wolfgang Hübner,et al.  Open-source image reconstruction of super-resolution structured illumination microscopy data in ImageJ , 2016, Nature Communications.

[8]  Bryant B. Chhun,et al.  Super-Resolution Video Microscopy of Live Cells by Structured Illumination , 2009, Nature Methods.

[9]  K. O’Holleran,et al.  High speed structured illumination microscopy in optically thick samples. , 2015, Methods.

[10]  T. Weil,et al.  3D Time-lapse Imaging and Quantification of Mitochondrial Dynamics , 2017, Scientific Reports.

[11]  Mats G. L. Gustafsson,et al.  A new approach to extended focus for high-speed high-resolution biological microscopy , 2006, SPIE BiOS.

[12]  Rainer Heintzmann,et al.  fastSIM: a practical implementation of fast structured illumination microscopy , 2015, Methods and applications in fluorescence.

[13]  Brian P. Mehl,et al.  Multifocus microscopy with precise color multi-phase diffractive optics applied in functional neuronal imaging. , 2016, Biomedical optics express.

[14]  M. Gustafsson.  Surpassing the lateral resolution limit by a factor of two using structured illumination microscopy , 2000, Journal of microscopy.

[15]  S. Hess,et al.  Three-dimensional sub–100 nm resolution fluorescence microscopy of thick samples , 2008, Nature Methods.

[16]  Alexander Y Katsov,et al.  fast multicolor 3 d imaging using aberration-corrected multifocus microscopy , 2012 .

[17]  Rainer Heintzmann,et al.  Laterally modulated excitation microscopy: improvement of resolution by using a diffraction grating , 1999, European Conference on Biomedical Optics.

[18]  M. Gustafsson,et al.  Three-dimensional resolution doubling in wide-field fluorescence microscopy by structured illumination. , 2008, Biophysical journal.

[19]  K. O’Holleran,et al.  Optimized approaches for optical sectioning and resolution enhancement in 2D structured illumination microscopy , 2014, Biomedical optics express.

[20]  Jianying Zhou,et al.  Fast structured illumination microscopy using rolling shutter cameras , 2016 .

[21]  T. Lukes,et al.  Combined multi-plane phase retrieval and super-resolution optical fluctuation imaging for 4D cell microscopy , 2018 .

[22]  S. Jakobs,et al.  Live-cell multiplane three-dimensional super-resolution optical fluctuation imaging , 2014, Nature Communications.

[23]  P. Xi,et al.  Fast, long-term, super-resolution imaging with Hessian structured illumination microscopy , 2018, Nature Biotechnology.