Abstract

Fluorescence microscopy is the standard imaging technique to investigate the structures and dynamics of living cells. However, increasing the spatial resolution comes at the cost of temporal resolution and vice versa. In addition, the number of images that can be taken in sufficiently high quality is limited by fluorescence bleaching. Hence, super-resolved imaging at several Hertz of low fluorescent biological samples is still a big challenge and, especially in structured illumination microscopy (SIM), is often visible as imaging artifacts. In this paper, we present a TIRF-SIM system based on scan-mirrors and a Michelson interferometer, which generates images at 110 nm spatial resolution and up to 8 Hz temporal resolution. High resolution becomes possible by optimizing the illumination interference contrast, even for low fluorescent, moving samples. We provide a framework and guidelines on how the modulation contrast, which depends on laser coherence, polarization, beam displacement or sample movements, can be mapped over the entire field of view. In addition, we characterize the influence of the signal-to-noise ratio and the Wiener filtering on the quality of reconstructed SIM images, both in real and frequency space. Our results are supported by theoretical descriptions containing the parameters leading to image artifacts. This study aims to help microscopists to better understand and adjust optical parameters for structured illumination, thereby leading to more trustworthy measurements and analyses of biological dynamics.

© 2020 Optical Society of America under the terms of the OSA Open Access Publishing Agreement

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2019 (1)

C. Billaudeau, Z. Yao, C. Cornilleau, R. Carballido-López, and A. Chastanet, “MreB Forms Subdiffraction Nanofilaments during Active Growth in Bacillus subtilis,” mBio 10(1), e01879-18 (2019).
[Crossref]

2018 (3)

X. Huang, J. Fan, L. Li, H. Liu, R. Wu, Y. Wu, L. Wei, H. Mao, A. Lal, P. Xi, L. Tang, Y. Zhang, Y. Liu, S. Tan, and L. Chen, “Fast, long-term, super-resolution imaging with Hessian structured illumination microscopy,” Nat. Biotechnol. 36(5), 451–459 (2018).
[Crossref]

Y. Chen, Y. Chen, R. Cao, W. Liu, D. Zhu, Z. Zhang, C. Kuang, and X. Liu, “Widefield and total internal reflection fluorescent structured illumination microscopy with scanning galvo mirrors structured illumination microscopy with scanning,” J. Biomed. Opt. 23(4), 046007 (2018).
[Crossref]

R. Förster, W. Müller, R. Richter, and R. Heintzmann, “Automated distinction of shearing and distortion artefacts in structured illumination microscopy,” Opt. Express 26(16), 20680 (2018).
[Crossref]

2017 (1)

J. Demmerle, C. Innocent, A. J. North, G. Ball, M. Müller, E. Miron, A. Matsuda, I. M. Dobbie, Y. Markaki, and L. Schermelleh, “Strategic and practical guidelines for successful structured illumination microscopy,” Nat. Protoc. 12(5), 988–1010 (2017).
[Crossref]

2016 (5)

S. J. Sahl, F. Balzarotti, J. Keller-Findeisen, M. Leutenegger, V. Westphal, A. Egner, F. Lavoie-Cardinal, A. Chmyrov, T. Grotjohann, and S. Jakobs, “Comment on “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 352(6285), 527 (2016).
[Crossref]

N. Chakrova, B. Rieger, and S. Stallinga, “Deconvolution methods for structured illumination microscopy,” J. Opt. Soc. Am. A 33(7), B12 (2016).
[Crossref]

F. Jünger, P. v. Olshausen, and A. Rohrbach, “Fast, label-free super-resolution live-cell imaging using rotating coherent scattering (ROCS) microscopy,” Sci. Rep. 6, 30393 (2016).
[Crossref]

R. Förster, K. Wicker, W. Müller, A. Jost, and R. Heintzmann, “Motion artefact detection in structured illumination microscopy for live cell imaging,” Opt. Express 24(19), 22121 (2016).
[Crossref]

L. J. Young, F. Ströhl, and C. F. Kaminski, “A Guide to Structured Illumination TIRF Microscopy at High Speed with Multiple Colors,” J. Visualized Exp. 111, 53988 (2016).
[Crossref]

2015 (4)

D. Li, L. Shao, B. C. Chen, X. Zhang, M. Zhang, B. Moses, D. E. Milkie, J. R. Beach, J. A. Hammer, M. Pasham, T. Kirchhausen, M. A. Baird, M. W. Davidson, P. Xu, and E. Betzig, “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 349(6251), aab3500 (2015).
[Crossref]

H.-W. Lu-Walther, M. Kielhorn, R. Förster, A. Jost, K. Wicker, and R. Heintzmann, “fastSIM: a practical implementation of fast structured illumination microscopy,” Methods Appl. Fluoresc. 3(1), 014001 (2015).
[Crossref]

F. Kohler and A. Rohrbach, “Surfing along filopodia: A particle transport revealed by molecular-scale fluctuation analyses,” Biophys. J 108, 2114–2125 (2015).
[Crossref]

G. Ball, J. Demmerle, R. Kaufmann, I. Davis, I. M. Dobbie, and L. Schermelleh, “SIMcheck: a Toolbox for Successful Super-resolution Structured Illumination Microscopy,” Sci. Rep. 5(1), 15915 (2015).
[Crossref]

2014 (3)

M. Brunstein, M. Teremetz, K. Hérault, C. Tourain, and M. Oheim, “Eliminating unwanted far-field excitation in objective-type TIRF. Part I. Identifying sources of nonevanescent excitation light,” Biophys. J. 106(5), 1020–1032 (2014).
[Crossref]

M. Brunstein, K. Hérault, and M. Oheim, “Eliminating unwanted far-field excitation in objective-type TIRF. Part II. Combined evanescent-wave excitation and supercritical-angle fluorescence detection improves optical sectioning,” Biophys. J. 106(5), 1044–1056 (2014).
[Crossref]

R. Förster, H.-W. Lu-Walther, A. Jost, M. Kielhorn, K. Wicker, and R. Heintzmann, “Simple structured illumination microscope setup with high acquisition speed by using a spatial light modulator,” Opt. Express 22(17), 20663 (2014).
[Crossref]

2013 (4)

2012 (1)

2009 (2)

L. M. Hirvonen, K. Wicker, O. Mandula, and R. Heintzmann, “Structured illumination microscopy of a living cell,” Eur. Biophys. J. 38(6), 807–812 (2009).
[Crossref]

P. Kner, B. B. Chhun, E. R. Griffis, L. Winoto, and M. G. L. Gustafsson, “Super-resolution video microscopy of live cells by structured illumination,” Nat. Methods 6(5), 339–342 (2009).
[Crossref]

2008 (1)

2000 (1)

M. G. Gustafsson, “Surpassing the lateral resolution limit by a factor of two using structured illumination microscopy,” J. Microsc. 198(2), 82–87 (2000).
[Crossref]

1984 (1)

D. Axelrod, T. P. Burghardt, and N. L. Thompson, “Total internal reflection fluorescence,” Annu. Rev. Biophys. Bioeng. 13(1), 247–268 (1984).
[Crossref]

Axelrod, D.

D. Axelrod, T. P. Burghardt, and N. L. Thompson, “Total internal reflection fluorescence,” Annu. Rev. Biophys. Bioeng. 13(1), 247–268 (1984).
[Crossref]

Baird, M. A.

D. Li, L. Shao, B. C. Chen, X. Zhang, M. Zhang, B. Moses, D. E. Milkie, J. R. Beach, J. A. Hammer, M. Pasham, T. Kirchhausen, M. A. Baird, M. W. Davidson, P. Xu, and E. Betzig, “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 349(6251), aab3500 (2015).
[Crossref]

Ball, G.

J. Demmerle, C. Innocent, A. J. North, G. Ball, M. Müller, E. Miron, A. Matsuda, I. M. Dobbie, Y. Markaki, and L. Schermelleh, “Strategic and practical guidelines for successful structured illumination microscopy,” Nat. Protoc. 12(5), 988–1010 (2017).
[Crossref]

G. Ball, J. Demmerle, R. Kaufmann, I. Davis, I. M. Dobbie, and L. Schermelleh, “SIMcheck: a Toolbox for Successful Super-resolution Structured Illumination Microscopy,” Sci. Rep. 5(1), 15915 (2015).
[Crossref]

Balzarotti, F.

S. J. Sahl, F. Balzarotti, J. Keller-Findeisen, M. Leutenegger, V. Westphal, A. Egner, F. Lavoie-Cardinal, A. Chmyrov, T. Grotjohann, and S. Jakobs, “Comment on “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 352(6285), 527 (2016).
[Crossref]

Beach, J. R.

D. Li, L. Shao, B. C. Chen, X. Zhang, M. Zhang, B. Moses, D. E. Milkie, J. R. Beach, J. A. Hammer, M. Pasham, T. Kirchhausen, M. A. Baird, M. W. Davidson, P. Xu, and E. Betzig, “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 349(6251), aab3500 (2015).
[Crossref]

Beck, M.

Best, G.

Betzig, E.

D. Li, L. Shao, B. C. Chen, X. Zhang, M. Zhang, B. Moses, D. E. Milkie, J. R. Beach, J. A. Hammer, M. Pasham, T. Kirchhausen, M. A. Baird, M. W. Davidson, P. Xu, and E. Betzig, “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 349(6251), aab3500 (2015).
[Crossref]

Billaudeau, C.

C. Billaudeau, Z. Yao, C. Cornilleau, R. Carballido-López, and A. Chastanet, “MreB Forms Subdiffraction Nanofilaments during Active Growth in Bacillus subtilis,” mBio 10(1), e01879-18 (2019).
[Crossref]

Brunstein, M.

M. Brunstein, K. Hérault, and M. Oheim, “Eliminating unwanted far-field excitation in objective-type TIRF. Part II. Combined evanescent-wave excitation and supercritical-angle fluorescence detection improves optical sectioning,” Biophys. J. 106(5), 1044–1056 (2014).
[Crossref]

M. Brunstein, M. Teremetz, K. Hérault, C. Tourain, and M. Oheim, “Eliminating unwanted far-field excitation in objective-type TIRF. Part I. Identifying sources of nonevanescent excitation light,” Biophys. J. 106(5), 1020–1032 (2014).
[Crossref]

M. Brunstein, K. Wicker, K. Hérault, R. Heintzmann, and M. Oheim, “Full-field dual-color 100-nm super-resolution imaging reveals organization and dynamics of mitochondrial and ER networks,” Opt. Express 21(22), 26162–26173 (2013).
[Crossref]

Burghardt, T. P.

D. Axelrod, T. P. Burghardt, and N. L. Thompson, “Total internal reflection fluorescence,” Annu. Rev. Biophys. Bioeng. 13(1), 247–268 (1984).
[Crossref]

Cao, R.

Y. Chen, Y. Chen, R. Cao, W. Liu, D. Zhu, Z. Zhang, C. Kuang, and X. Liu, “Widefield and total internal reflection fluorescent structured illumination microscopy with scanning galvo mirrors structured illumination microscopy with scanning,” J. Biomed. Opt. 23(4), 046007 (2018).
[Crossref]

Carballido-López, R.

C. Billaudeau, Z. Yao, C. Cornilleau, R. Carballido-López, and A. Chastanet, “MreB Forms Subdiffraction Nanofilaments during Active Growth in Bacillus subtilis,” mBio 10(1), e01879-18 (2019).
[Crossref]

Chakrova, N.

Chastanet, A.

C. Billaudeau, Z. Yao, C. Cornilleau, R. Carballido-López, and A. Chastanet, “MreB Forms Subdiffraction Nanofilaments during Active Growth in Bacillus subtilis,” mBio 10(1), e01879-18 (2019).
[Crossref]

Chen, B. C.

D. Li, L. Shao, B. C. Chen, X. Zhang, M. Zhang, B. Moses, D. E. Milkie, J. R. Beach, J. A. Hammer, M. Pasham, T. Kirchhausen, M. A. Baird, M. W. Davidson, P. Xu, and E. Betzig, “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 349(6251), aab3500 (2015).
[Crossref]

Chen, L.

X. Huang, J. Fan, L. Li, H. Liu, R. Wu, Y. Wu, L. Wei, H. Mao, A. Lal, P. Xi, L. Tang, Y. Zhang, Y. Liu, S. Tan, and L. Chen, “Fast, long-term, super-resolution imaging with Hessian structured illumination microscopy,” Nat. Biotechnol. 36(5), 451–459 (2018).
[Crossref]

Chen, Y.

Y. Chen, Y. Chen, R. Cao, W. Liu, D. Zhu, Z. Zhang, C. Kuang, and X. Liu, “Widefield and total internal reflection fluorescent structured illumination microscopy with scanning galvo mirrors structured illumination microscopy with scanning,” J. Biomed. Opt. 23(4), 046007 (2018).
[Crossref]

Y. Chen, Y. Chen, R. Cao, W. Liu, D. Zhu, Z. Zhang, C. Kuang, and X. Liu, “Widefield and total internal reflection fluorescent structured illumination microscopy with scanning galvo mirrors structured illumination microscopy with scanning,” J. Biomed. Opt. 23(4), 046007 (2018).
[Crossref]

Chhun, B. B.

P. Kner, B. B. Chhun, E. R. Griffis, L. Winoto, and M. G. L. Gustafsson, “Super-resolution video microscopy of live cells by structured illumination,” Nat. Methods 6(5), 339–342 (2009).
[Crossref]

Chmyrov, A.

S. J. Sahl, F. Balzarotti, J. Keller-Findeisen, M. Leutenegger, V. Westphal, A. Egner, F. Lavoie-Cardinal, A. Chmyrov, T. Grotjohann, and S. Jakobs, “Comment on “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 352(6285), 527 (2016).
[Crossref]

Cornilleau, C.

C. Billaudeau, Z. Yao, C. Cornilleau, R. Carballido-López, and A. Chastanet, “MreB Forms Subdiffraction Nanofilaments during Active Growth in Bacillus subtilis,” mBio 10(1), e01879-18 (2019).
[Crossref]

Davidson, M. W.

D. Li, L. Shao, B. C. Chen, X. Zhang, M. Zhang, B. Moses, D. E. Milkie, J. R. Beach, J. A. Hammer, M. Pasham, T. Kirchhausen, M. A. Baird, M. W. Davidson, P. Xu, and E. Betzig, “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 349(6251), aab3500 (2015).
[Crossref]

Davis, I.

G. Ball, J. Demmerle, R. Kaufmann, I. Davis, I. M. Dobbie, and L. Schermelleh, “SIMcheck: a Toolbox for Successful Super-resolution Structured Illumination Microscopy,” Sci. Rep. 5(1), 15915 (2015).
[Crossref]

Defeu Soufo, H. J.

P. V. Olshausen, H. J. Defeu Soufo, K. Wicker, R. Heintzmann, P. L. Graumann, and A. Rohrbach, “Superresolution imaging of dynamic MreB filaments in B. subtilis - A multiple-motor-driven transport?” Biophys. J. 105(5), 1171–1181 (2013).
[Crossref]

Demmerle, J.

J. Demmerle, C. Innocent, A. J. North, G. Ball, M. Müller, E. Miron, A. Matsuda, I. M. Dobbie, Y. Markaki, and L. Schermelleh, “Strategic and practical guidelines for successful structured illumination microscopy,” Nat. Protoc. 12(5), 988–1010 (2017).
[Crossref]

G. Ball, J. Demmerle, R. Kaufmann, I. Davis, I. M. Dobbie, and L. Schermelleh, “SIMcheck: a Toolbox for Successful Super-resolution Structured Illumination Microscopy,” Sci. Rep. 5(1), 15915 (2015).
[Crossref]

Dobbie, I. M.

J. Demmerle, C. Innocent, A. J. North, G. Ball, M. Müller, E. Miron, A. Matsuda, I. M. Dobbie, Y. Markaki, and L. Schermelleh, “Strategic and practical guidelines for successful structured illumination microscopy,” Nat. Protoc. 12(5), 988–1010 (2017).
[Crossref]

G. Ball, J. Demmerle, R. Kaufmann, I. Davis, I. M. Dobbie, and L. Schermelleh, “SIMcheck: a Toolbox for Successful Super-resolution Structured Illumination Microscopy,” Sci. Rep. 5(1), 15915 (2015).
[Crossref]

Egner, A.

S. J. Sahl, F. Balzarotti, J. Keller-Findeisen, M. Leutenegger, V. Westphal, A. Egner, F. Lavoie-Cardinal, A. Chmyrov, T. Grotjohann, and S. Jakobs, “Comment on “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 352(6285), 527 (2016).
[Crossref]

Fan, J.

X. Huang, J. Fan, L. Li, H. Liu, R. Wu, Y. Wu, L. Wei, H. Mao, A. Lal, P. Xi, L. Tang, Y. Zhang, Y. Liu, S. Tan, and L. Chen, “Fast, long-term, super-resolution imaging with Hessian structured illumination microscopy,” Nat. Biotechnol. 36(5), 451–459 (2018).
[Crossref]

Fiolka, R.

Fliegel, K.

J. Pospíšil, K. Fliegel, and M. Klíma, “Analysis of image reconstruction artifacts in structured illumination microscopy,” in Applications of Digital Image Processing XL, A. G. Tescher, ed. (SPIE, 2017), Vol. 10396, p. 110.

Förster, R.

Graumann, P. L.

P. V. Olshausen, H. J. Defeu Soufo, K. Wicker, R. Heintzmann, P. L. Graumann, and A. Rohrbach, “Superresolution imaging of dynamic MreB filaments in B. subtilis - A multiple-motor-driven transport?” Biophys. J. 105(5), 1171–1181 (2013).
[Crossref]

Griffis, E. R.

P. Kner, B. B. Chhun, E. R. Griffis, L. Winoto, and M. G. L. Gustafsson, “Super-resolution video microscopy of live cells by structured illumination,” Nat. Methods 6(5), 339–342 (2009).
[Crossref]

Grotjohann, T.

S. J. Sahl, F. Balzarotti, J. Keller-Findeisen, M. Leutenegger, V. Westphal, A. Egner, F. Lavoie-Cardinal, A. Chmyrov, T. Grotjohann, and S. Jakobs, “Comment on “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 352(6285), 527 (2016).
[Crossref]

Gustafsson, M. G.

M. G. Gustafsson, “Surpassing the lateral resolution limit by a factor of two using structured illumination microscopy,” J. Microsc. 198(2), 82–87 (2000).
[Crossref]

Gustafsson, M. G. L.

P. Kner, B. B. Chhun, E. R. Griffis, L. Winoto, and M. G. L. Gustafsson, “Super-resolution video microscopy of live cells by structured illumination,” Nat. Methods 6(5), 339–342 (2009).
[Crossref]

Hammer, J. A.

D. Li, L. Shao, B. C. Chen, X. Zhang, M. Zhang, B. Moses, D. E. Milkie, J. R. Beach, J. A. Hammer, M. Pasham, T. Kirchhausen, M. A. Baird, M. W. Davidson, P. Xu, and E. Betzig, “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 349(6251), aab3500 (2015).
[Crossref]

Heintzmann, R.

R. Förster, W. Müller, R. Richter, and R. Heintzmann, “Automated distinction of shearing and distortion artefacts in structured illumination microscopy,” Opt. Express 26(16), 20680 (2018).
[Crossref]

R. Förster, K. Wicker, W. Müller, A. Jost, and R. Heintzmann, “Motion artefact detection in structured illumination microscopy for live cell imaging,” Opt. Express 24(19), 22121 (2016).
[Crossref]

H.-W. Lu-Walther, M. Kielhorn, R. Förster, A. Jost, K. Wicker, and R. Heintzmann, “fastSIM: a practical implementation of fast structured illumination microscopy,” Methods Appl. Fluoresc. 3(1), 014001 (2015).
[Crossref]

R. Förster, H.-W. Lu-Walther, A. Jost, M. Kielhorn, K. Wicker, and R. Heintzmann, “Simple structured illumination microscope setup with high acquisition speed by using a spatial light modulator,” Opt. Express 22(17), 20663 (2014).
[Crossref]

M. Brunstein, K. Wicker, K. Hérault, R. Heintzmann, and M. Oheim, “Full-field dual-color 100-nm super-resolution imaging reveals organization and dynamics of mitochondrial and ER networks,” Opt. Express 21(22), 26162–26173 (2013).
[Crossref]

P. V. Olshausen, H. J. Defeu Soufo, K. Wicker, R. Heintzmann, P. L. Graumann, and A. Rohrbach, “Superresolution imaging of dynamic MreB filaments in B. subtilis - A multiple-motor-driven transport?” Biophys. J. 105(5), 1171–1181 (2013).
[Crossref]

K. Wicker, O. Mandula, G. Best, R. Fiolka, and R. Heintzmann, “Phase optimisation for structured illumination microscopy,” Opt. Express 21(2), 2032 (2013).
[Crossref]

L. M. Hirvonen, K. Wicker, O. Mandula, and R. Heintzmann, “Structured illumination microscopy of a living cell,” Eur. Biophys. J. 38(6), 807–812 (2009).
[Crossref]

Hérault, K.

M. Brunstein, K. Hérault, and M. Oheim, “Eliminating unwanted far-field excitation in objective-type TIRF. Part II. Combined evanescent-wave excitation and supercritical-angle fluorescence detection improves optical sectioning,” Biophys. J. 106(5), 1044–1056 (2014).
[Crossref]

M. Brunstein, M. Teremetz, K. Hérault, C. Tourain, and M. Oheim, “Eliminating unwanted far-field excitation in objective-type TIRF. Part I. Identifying sources of nonevanescent excitation light,” Biophys. J. 106(5), 1020–1032 (2014).
[Crossref]

M. Brunstein, K. Wicker, K. Hérault, R. Heintzmann, and M. Oheim, “Full-field dual-color 100-nm super-resolution imaging reveals organization and dynamics of mitochondrial and ER networks,” Opt. Express 21(22), 26162–26173 (2013).
[Crossref]

Hirvonen, L. M.

L. M. Hirvonen, K. Wicker, O. Mandula, and R. Heintzmann, “Structured illumination microscopy of a living cell,” Eur. Biophys. J. 38(6), 807–812 (2009).
[Crossref]

Holleran, K. O.

Huang, X.

X. Huang, J. Fan, L. Li, H. Liu, R. Wu, Y. Wu, L. Wei, H. Mao, A. Lal, P. Xi, L. Tang, Y. Zhang, Y. Liu, S. Tan, and L. Chen, “Fast, long-term, super-resolution imaging with Hessian structured illumination microscopy,” Nat. Biotechnol. 36(5), 451–459 (2018).
[Crossref]

Innocent, C.

J. Demmerle, C. Innocent, A. J. North, G. Ball, M. Müller, E. Miron, A. Matsuda, I. M. Dobbie, Y. Markaki, and L. Schermelleh, “Strategic and practical guidelines for successful structured illumination microscopy,” Nat. Protoc. 12(5), 988–1010 (2017).
[Crossref]

Jakobs, S.

S. J. Sahl, F. Balzarotti, J. Keller-Findeisen, M. Leutenegger, V. Westphal, A. Egner, F. Lavoie-Cardinal, A. Chmyrov, T. Grotjohann, and S. Jakobs, “Comment on “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 352(6285), 527 (2016).
[Crossref]

Jost, A.

Jünger, F.

F. Jünger, P. v. Olshausen, and A. Rohrbach, “Fast, label-free super-resolution live-cell imaging using rotating coherent scattering (ROCS) microscopy,” Sci. Rep. 6, 30393 (2016).
[Crossref]

F. Jünger and A. Rohrbach, “Strong cytoskeleton activity on millisecond timescales upon particle binding revealed by ROCS microscopy,” Cytoskeleton (2018).

Kaminski, C. F.

L. J. Young, F. Ströhl, and C. F. Kaminski, “A Guide to Structured Illumination TIRF Microscopy at High Speed with Multiple Colors,” J. Visualized Exp. 111, 53988 (2016).
[Crossref]

Kaufmann, R.

G. Ball, J. Demmerle, R. Kaufmann, I. Davis, I. M. Dobbie, and L. Schermelleh, “SIMcheck: a Toolbox for Successful Super-resolution Structured Illumination Microscopy,” Sci. Rep. 5(1), 15915 (2015).
[Crossref]

Keller-Findeisen, J.

S. J. Sahl, F. Balzarotti, J. Keller-Findeisen, M. Leutenegger, V. Westphal, A. Egner, F. Lavoie-Cardinal, A. Chmyrov, T. Grotjohann, and S. Jakobs, “Comment on “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 352(6285), 527 (2016).
[Crossref]

Kielhorn, M.

H.-W. Lu-Walther, M. Kielhorn, R. Förster, A. Jost, K. Wicker, and R. Heintzmann, “fastSIM: a practical implementation of fast structured illumination microscopy,” Methods Appl. Fluoresc. 3(1), 014001 (2015).
[Crossref]

R. Förster, H.-W. Lu-Walther, A. Jost, M. Kielhorn, K. Wicker, and R. Heintzmann, “Simple structured illumination microscope setup with high acquisition speed by using a spatial light modulator,” Opt. Express 22(17), 20663 (2014).
[Crossref]

Kirchhausen, T.

D. Li, L. Shao, B. C. Chen, X. Zhang, M. Zhang, B. Moses, D. E. Milkie, J. R. Beach, J. A. Hammer, M. Pasham, T. Kirchhausen, M. A. Baird, M. W. Davidson, P. Xu, and E. Betzig, “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 349(6251), aab3500 (2015).
[Crossref]

Klíma, M.

J. Pospíšil, K. Fliegel, and M. Klíma, “Analysis of image reconstruction artifacts in structured illumination microscopy,” in Applications of Digital Image Processing XL, A. G. Tescher, ed. (SPIE, 2017), Vol. 10396, p. 110.

Kner, P.

P. Kner, B. B. Chhun, E. R. Griffis, L. Winoto, and M. G. L. Gustafsson, “Super-resolution video microscopy of live cells by structured illumination,” Nat. Methods 6(5), 339–342 (2009).
[Crossref]

Kohler, F.

F. Kohler and A. Rohrbach, “Surfing along filopodia: A particle transport revealed by molecular-scale fluctuation analyses,” Biophys. J 108, 2114–2125 (2015).
[Crossref]

Kuang, C.

Y. Chen, Y. Chen, R. Cao, W. Liu, D. Zhu, Z. Zhang, C. Kuang, and X. Liu, “Widefield and total internal reflection fluorescent structured illumination microscopy with scanning galvo mirrors structured illumination microscopy with scanning,” J. Biomed. Opt. 23(4), 046007 (2018).
[Crossref]

Lal, A.

X. Huang, J. Fan, L. Li, H. Liu, R. Wu, Y. Wu, L. Wei, H. Mao, A. Lal, P. Xi, L. Tang, Y. Zhang, Y. Liu, S. Tan, and L. Chen, “Fast, long-term, super-resolution imaging with Hessian structured illumination microscopy,” Nat. Biotechnol. 36(5), 451–459 (2018).
[Crossref]

Lavoie-Cardinal, F.

S. J. Sahl, F. Balzarotti, J. Keller-Findeisen, M. Leutenegger, V. Westphal, A. Egner, F. Lavoie-Cardinal, A. Chmyrov, T. Grotjohann, and S. Jakobs, “Comment on “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 352(6285), 527 (2016).
[Crossref]

Leutenegger, M.

S. J. Sahl, F. Balzarotti, J. Keller-Findeisen, M. Leutenegger, V. Westphal, A. Egner, F. Lavoie-Cardinal, A. Chmyrov, T. Grotjohann, and S. Jakobs, “Comment on “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 352(6285), 527 (2016).
[Crossref]

Li, D.

D. Li, L. Shao, B. C. Chen, X. Zhang, M. Zhang, B. Moses, D. E. Milkie, J. R. Beach, J. A. Hammer, M. Pasham, T. Kirchhausen, M. A. Baird, M. W. Davidson, P. Xu, and E. Betzig, “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 349(6251), aab3500 (2015).
[Crossref]

Li, L.

X. Huang, J. Fan, L. Li, H. Liu, R. Wu, Y. Wu, L. Wei, H. Mao, A. Lal, P. Xi, L. Tang, Y. Zhang, Y. Liu, S. Tan, and L. Chen, “Fast, long-term, super-resolution imaging with Hessian structured illumination microscopy,” Nat. Biotechnol. 36(5), 451–459 (2018).
[Crossref]

Liu, H.

X. Huang, J. Fan, L. Li, H. Liu, R. Wu, Y. Wu, L. Wei, H. Mao, A. Lal, P. Xi, L. Tang, Y. Zhang, Y. Liu, S. Tan, and L. Chen, “Fast, long-term, super-resolution imaging with Hessian structured illumination microscopy,” Nat. Biotechnol. 36(5), 451–459 (2018).
[Crossref]

Liu, W.

Y. Chen, Y. Chen, R. Cao, W. Liu, D. Zhu, Z. Zhang, C. Kuang, and X. Liu, “Widefield and total internal reflection fluorescent structured illumination microscopy with scanning galvo mirrors structured illumination microscopy with scanning,” J. Biomed. Opt. 23(4), 046007 (2018).
[Crossref]

Liu, X.

Y. Chen, Y. Chen, R. Cao, W. Liu, D. Zhu, Z. Zhang, C. Kuang, and X. Liu, “Widefield and total internal reflection fluorescent structured illumination microscopy with scanning galvo mirrors structured illumination microscopy with scanning,” J. Biomed. Opt. 23(4), 046007 (2018).
[Crossref]

Liu, Y.

X. Huang, J. Fan, L. Li, H. Liu, R. Wu, Y. Wu, L. Wei, H. Mao, A. Lal, P. Xi, L. Tang, Y. Zhang, Y. Liu, S. Tan, and L. Chen, “Fast, long-term, super-resolution imaging with Hessian structured illumination microscopy,” Nat. Biotechnol. 36(5), 451–459 (2018).
[Crossref]

Lu-Walther, H.-W.

H.-W. Lu-Walther, M. Kielhorn, R. Förster, A. Jost, K. Wicker, and R. Heintzmann, “fastSIM: a practical implementation of fast structured illumination microscopy,” Methods Appl. Fluoresc. 3(1), 014001 (2015).
[Crossref]

R. Förster, H.-W. Lu-Walther, A. Jost, M. Kielhorn, K. Wicker, and R. Heintzmann, “Simple structured illumination microscope setup with high acquisition speed by using a spatial light modulator,” Opt. Express 22(17), 20663 (2014).
[Crossref]

Mandula, O.

K. Wicker, O. Mandula, G. Best, R. Fiolka, and R. Heintzmann, “Phase optimisation for structured illumination microscopy,” Opt. Express 21(2), 2032 (2013).
[Crossref]

L. M. Hirvonen, K. Wicker, O. Mandula, and R. Heintzmann, “Structured illumination microscopy of a living cell,” Eur. Biophys. J. 38(6), 807–812 (2009).
[Crossref]

Mao, H.

X. Huang, J. Fan, L. Li, H. Liu, R. Wu, Y. Wu, L. Wei, H. Mao, A. Lal, P. Xi, L. Tang, Y. Zhang, Y. Liu, S. Tan, and L. Chen, “Fast, long-term, super-resolution imaging with Hessian structured illumination microscopy,” Nat. Biotechnol. 36(5), 451–459 (2018).
[Crossref]

Markaki, Y.

J. Demmerle, C. Innocent, A. J. North, G. Ball, M. Müller, E. Miron, A. Matsuda, I. M. Dobbie, Y. Markaki, and L. Schermelleh, “Strategic and practical guidelines for successful structured illumination microscopy,” Nat. Protoc. 12(5), 988–1010 (2017).
[Crossref]

Matsuda, A.

J. Demmerle, C. Innocent, A. J. North, G. Ball, M. Müller, E. Miron, A. Matsuda, I. M. Dobbie, Y. Markaki, and L. Schermelleh, “Strategic and practical guidelines for successful structured illumination microscopy,” Nat. Protoc. 12(5), 988–1010 (2017).
[Crossref]

Milkie, D. E.

D. Li, L. Shao, B. C. Chen, X. Zhang, M. Zhang, B. Moses, D. E. Milkie, J. R. Beach, J. A. Hammer, M. Pasham, T. Kirchhausen, M. A. Baird, M. W. Davidson, P. Xu, and E. Betzig, “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 349(6251), aab3500 (2015).
[Crossref]

Miron, E.

J. Demmerle, C. Innocent, A. J. North, G. Ball, M. Müller, E. Miron, A. Matsuda, I. M. Dobbie, Y. Markaki, and L. Schermelleh, “Strategic and practical guidelines for successful structured illumination microscopy,” Nat. Protoc. 12(5), 988–1010 (2017).
[Crossref]

Moses, B.

D. Li, L. Shao, B. C. Chen, X. Zhang, M. Zhang, B. Moses, D. E. Milkie, J. R. Beach, J. A. Hammer, M. Pasham, T. Kirchhausen, M. A. Baird, M. W. Davidson, P. Xu, and E. Betzig, “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 349(6251), aab3500 (2015).
[Crossref]

Müller, M.

J. Demmerle, C. Innocent, A. J. North, G. Ball, M. Müller, E. Miron, A. Matsuda, I. M. Dobbie, Y. Markaki, and L. Schermelleh, “Strategic and practical guidelines for successful structured illumination microscopy,” Nat. Protoc. 12(5), 988–1010 (2017).
[Crossref]

Müller, W.

North, A. J.

J. Demmerle, C. Innocent, A. J. North, G. Ball, M. Müller, E. Miron, A. Matsuda, I. M. Dobbie, Y. Markaki, and L. Schermelleh, “Strategic and practical guidelines for successful structured illumination microscopy,” Nat. Protoc. 12(5), 988–1010 (2017).
[Crossref]

O’Holleran, K.

Oheim, M.

M. Brunstein, K. Hérault, and M. Oheim, “Eliminating unwanted far-field excitation in objective-type TIRF. Part II. Combined evanescent-wave excitation and supercritical-angle fluorescence detection improves optical sectioning,” Biophys. J. 106(5), 1044–1056 (2014).
[Crossref]

M. Brunstein, M. Teremetz, K. Hérault, C. Tourain, and M. Oheim, “Eliminating unwanted far-field excitation in objective-type TIRF. Part I. Identifying sources of nonevanescent excitation light,” Biophys. J. 106(5), 1020–1032 (2014).
[Crossref]

M. Brunstein, K. Wicker, K. Hérault, R. Heintzmann, and M. Oheim, “Full-field dual-color 100-nm super-resolution imaging reveals organization and dynamics of mitochondrial and ER networks,” Opt. Express 21(22), 26162–26173 (2013).
[Crossref]

Olshausen, P. v.

F. Jünger, P. v. Olshausen, and A. Rohrbach, “Fast, label-free super-resolution live-cell imaging using rotating coherent scattering (ROCS) microscopy,” Sci. Rep. 6, 30393 (2016).
[Crossref]

P. V. Olshausen, H. J. Defeu Soufo, K. Wicker, R. Heintzmann, P. L. Graumann, and A. Rohrbach, “Superresolution imaging of dynamic MreB filaments in B. subtilis - A multiple-motor-driven transport?” Biophys. J. 105(5), 1171–1181 (2013).
[Crossref]

Pasham, M.

D. Li, L. Shao, B. C. Chen, X. Zhang, M. Zhang, B. Moses, D. E. Milkie, J. R. Beach, J. A. Hammer, M. Pasham, T. Kirchhausen, M. A. Baird, M. W. Davidson, P. Xu, and E. Betzig, “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 349(6251), aab3500 (2015).
[Crossref]

Pospíšil, J.

J. Pospíšil, K. Fliegel, and M. Klíma, “Analysis of image reconstruction artifacts in structured illumination microscopy,” in Applications of Digital Image Processing XL, A. G. Tescher, ed. (SPIE, 2017), Vol. 10396, p. 110.

Richter, R.

Rieger, B.

Rohrbach, A.

F. Jünger, P. v. Olshausen, and A. Rohrbach, “Fast, label-free super-resolution live-cell imaging using rotating coherent scattering (ROCS) microscopy,” Sci. Rep. 6, 30393 (2016).
[Crossref]

F. Kohler and A. Rohrbach, “Surfing along filopodia: A particle transport revealed by molecular-scale fluctuation analyses,” Biophys. J 108, 2114–2125 (2015).
[Crossref]

P. V. Olshausen, H. J. Defeu Soufo, K. Wicker, R. Heintzmann, P. L. Graumann, and A. Rohrbach, “Superresolution imaging of dynamic MreB filaments in B. subtilis - A multiple-motor-driven transport?” Biophys. J. 105(5), 1171–1181 (2013).
[Crossref]

F. Jünger and A. Rohrbach, “Strong cytoskeleton activity on millisecond timescales upon particle binding revealed by ROCS microscopy,” Cytoskeleton (2018).

Sahl, S. J.

S. J. Sahl, F. Balzarotti, J. Keller-Findeisen, M. Leutenegger, V. Westphal, A. Egner, F. Lavoie-Cardinal, A. Chmyrov, T. Grotjohann, and S. Jakobs, “Comment on “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 352(6285), 527 (2016).
[Crossref]

Schermelleh, L.

J. Demmerle, C. Innocent, A. J. North, G. Ball, M. Müller, E. Miron, A. Matsuda, I. M. Dobbie, Y. Markaki, and L. Schermelleh, “Strategic and practical guidelines for successful structured illumination microscopy,” Nat. Protoc. 12(5), 988–1010 (2017).
[Crossref]

G. Ball, J. Demmerle, R. Kaufmann, I. Davis, I. M. Dobbie, and L. Schermelleh, “SIMcheck: a Toolbox for Successful Super-resolution Structured Illumination Microscopy,” Sci. Rep. 5(1), 15915 (2015).
[Crossref]

Shao, L.

D. Li, L. Shao, B. C. Chen, X. Zhang, M. Zhang, B. Moses, D. E. Milkie, J. R. Beach, J. A. Hammer, M. Pasham, T. Kirchhausen, M. A. Baird, M. W. Davidson, P. Xu, and E. Betzig, “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 349(6251), aab3500 (2015).
[Crossref]

Shaw, M.

Stallinga, S.

Stemmer, A.

Ströhl, F.

L. J. Young, F. Ströhl, and C. F. Kaminski, “A Guide to Structured Illumination TIRF Microscopy at High Speed with Multiple Colors,” J. Visualized Exp. 111, 53988 (2016).
[Crossref]

Tan, S.

X. Huang, J. Fan, L. Li, H. Liu, R. Wu, Y. Wu, L. Wei, H. Mao, A. Lal, P. Xi, L. Tang, Y. Zhang, Y. Liu, S. Tan, and L. Chen, “Fast, long-term, super-resolution imaging with Hessian structured illumination microscopy,” Nat. Biotechnol. 36(5), 451–459 (2018).
[Crossref]

Tang, L.

X. Huang, J. Fan, L. Li, H. Liu, R. Wu, Y. Wu, L. Wei, H. Mao, A. Lal, P. Xi, L. Tang, Y. Zhang, Y. Liu, S. Tan, and L. Chen, “Fast, long-term, super-resolution imaging with Hessian structured illumination microscopy,” Nat. Biotechnol. 36(5), 451–459 (2018).
[Crossref]

Teremetz, M.

M. Brunstein, M. Teremetz, K. Hérault, C. Tourain, and M. Oheim, “Eliminating unwanted far-field excitation in objective-type TIRF. Part I. Identifying sources of nonevanescent excitation light,” Biophys. J. 106(5), 1020–1032 (2014).
[Crossref]

Thompson, N. L.

D. Axelrod, T. P. Burghardt, and N. L. Thompson, “Total internal reflection fluorescence,” Annu. Rev. Biophys. Bioeng. 13(1), 247–268 (1984).
[Crossref]

Tourain, C.

M. Brunstein, M. Teremetz, K. Hérault, C. Tourain, and M. Oheim, “Eliminating unwanted far-field excitation in objective-type TIRF. Part I. Identifying sources of nonevanescent excitation light,” Biophys. J. 106(5), 1020–1032 (2014).
[Crossref]

von Olshausen, P.

P. von Olshausen, “Total internal reflection microscopy: super-resolution imaging of bacterial dynamics and dark field imaging,” (2012).

Wei, L.

X. Huang, J. Fan, L. Li, H. Liu, R. Wu, Y. Wu, L. Wei, H. Mao, A. Lal, P. Xi, L. Tang, Y. Zhang, Y. Liu, S. Tan, and L. Chen, “Fast, long-term, super-resolution imaging with Hessian structured illumination microscopy,” Nat. Biotechnol. 36(5), 451–459 (2018).
[Crossref]

Westphal, V.

S. J. Sahl, F. Balzarotti, J. Keller-Findeisen, M. Leutenegger, V. Westphal, A. Egner, F. Lavoie-Cardinal, A. Chmyrov, T. Grotjohann, and S. Jakobs, “Comment on “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 352(6285), 527 (2016).
[Crossref]

Wicker, K.

R. Förster, K. Wicker, W. Müller, A. Jost, and R. Heintzmann, “Motion artefact detection in structured illumination microscopy for live cell imaging,” Opt. Express 24(19), 22121 (2016).
[Crossref]

H.-W. Lu-Walther, M. Kielhorn, R. Förster, A. Jost, K. Wicker, and R. Heintzmann, “fastSIM: a practical implementation of fast structured illumination microscopy,” Methods Appl. Fluoresc. 3(1), 014001 (2015).
[Crossref]

R. Förster, H.-W. Lu-Walther, A. Jost, M. Kielhorn, K. Wicker, and R. Heintzmann, “Simple structured illumination microscope setup with high acquisition speed by using a spatial light modulator,” Opt. Express 22(17), 20663 (2014).
[Crossref]

M. Brunstein, K. Wicker, K. Hérault, R. Heintzmann, and M. Oheim, “Full-field dual-color 100-nm super-resolution imaging reveals organization and dynamics of mitochondrial and ER networks,” Opt. Express 21(22), 26162–26173 (2013).
[Crossref]

P. V. Olshausen, H. J. Defeu Soufo, K. Wicker, R. Heintzmann, P. L. Graumann, and A. Rohrbach, “Superresolution imaging of dynamic MreB filaments in B. subtilis - A multiple-motor-driven transport?” Biophys. J. 105(5), 1171–1181 (2013).
[Crossref]

K. Wicker, “Non-iterative determination of pattern phase in structured illumination microscopy using auto-correlations in Fourier space,” Opt. Express 21(21), 24692 (2013).
[Crossref]

K. Wicker, O. Mandula, G. Best, R. Fiolka, and R. Heintzmann, “Phase optimisation for structured illumination microscopy,” Opt. Express 21(2), 2032 (2013).
[Crossref]

L. M. Hirvonen, K. Wicker, O. Mandula, and R. Heintzmann, “Structured illumination microscopy of a living cell,” Eur. Biophys. J. 38(6), 807–812 (2009).
[Crossref]

Winoto, L.

P. Kner, B. B. Chhun, E. R. Griffis, L. Winoto, and M. G. L. Gustafsson, “Super-resolution video microscopy of live cells by structured illumination,” Nat. Methods 6(5), 339–342 (2009).
[Crossref]

Wu, R.

X. Huang, J. Fan, L. Li, H. Liu, R. Wu, Y. Wu, L. Wei, H. Mao, A. Lal, P. Xi, L. Tang, Y. Zhang, Y. Liu, S. Tan, and L. Chen, “Fast, long-term, super-resolution imaging with Hessian structured illumination microscopy,” Nat. Biotechnol. 36(5), 451–459 (2018).
[Crossref]

Wu, Y.

X. Huang, J. Fan, L. Li, H. Liu, R. Wu, Y. Wu, L. Wei, H. Mao, A. Lal, P. Xi, L. Tang, Y. Zhang, Y. Liu, S. Tan, and L. Chen, “Fast, long-term, super-resolution imaging with Hessian structured illumination microscopy,” Nat. Biotechnol. 36(5), 451–459 (2018).
[Crossref]

Xi, P.

X. Huang, J. Fan, L. Li, H. Liu, R. Wu, Y. Wu, L. Wei, H. Mao, A. Lal, P. Xi, L. Tang, Y. Zhang, Y. Liu, S. Tan, and L. Chen, “Fast, long-term, super-resolution imaging with Hessian structured illumination microscopy,” Nat. Biotechnol. 36(5), 451–459 (2018).
[Crossref]

Xu, P.

D. Li, L. Shao, B. C. Chen, X. Zhang, M. Zhang, B. Moses, D. E. Milkie, J. R. Beach, J. A. Hammer, M. Pasham, T. Kirchhausen, M. A. Baird, M. W. Davidson, P. Xu, and E. Betzig, “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 349(6251), aab3500 (2015).
[Crossref]

Yao, Z.

C. Billaudeau, Z. Yao, C. Cornilleau, R. Carballido-López, and A. Chastanet, “MreB Forms Subdiffraction Nanofilaments during Active Growth in Bacillus subtilis,” mBio 10(1), e01879-18 (2019).
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Young, L. J.

L. J. Young, F. Ströhl, and C. F. Kaminski, “A Guide to Structured Illumination TIRF Microscopy at High Speed with Multiple Colors,” J. Visualized Exp. 111, 53988 (2016).
[Crossref]

Zhang, M.

D. Li, L. Shao, B. C. Chen, X. Zhang, M. Zhang, B. Moses, D. E. Milkie, J. R. Beach, J. A. Hammer, M. Pasham, T. Kirchhausen, M. A. Baird, M. W. Davidson, P. Xu, and E. Betzig, “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 349(6251), aab3500 (2015).
[Crossref]

Zhang, X.

D. Li, L. Shao, B. C. Chen, X. Zhang, M. Zhang, B. Moses, D. E. Milkie, J. R. Beach, J. A. Hammer, M. Pasham, T. Kirchhausen, M. A. Baird, M. W. Davidson, P. Xu, and E. Betzig, “Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics,” Science 349(6251), aab3500 (2015).
[Crossref]

Zhang, Y.

X. Huang, J. Fan, L. Li, H. Liu, R. Wu, Y. Wu, L. Wei, H. Mao, A. Lal, P. Xi, L. Tang, Y. Zhang, Y. Liu, S. Tan, and L. Chen, “Fast, long-term, super-resolution imaging with Hessian structured illumination microscopy,” Nat. Biotechnol. 36(5), 451–459 (2018).
[Crossref]

Zhang, Z.

Y. Chen, Y. Chen, R. Cao, W. Liu, D. Zhu, Z. Zhang, C. Kuang, and X. Liu, “Widefield and total internal reflection fluorescent structured illumination microscopy with scanning galvo mirrors structured illumination microscopy with scanning,” J. Biomed. Opt. 23(4), 046007 (2018).
[Crossref]

Zhu, D.

Y. Chen, Y. Chen, R. Cao, W. Liu, D. Zhu, Z. Zhang, C. Kuang, and X. Liu, “Widefield and total internal reflection fluorescent structured illumination microscopy with scanning galvo mirrors structured illumination microscopy with scanning,” J. Biomed. Opt. 23(4), 046007 (2018).
[Crossref]

Annu. Rev. Biophys. Bioeng. (1)

D. Axelrod, T. P. Burghardt, and N. L. Thompson, “Total internal reflection fluorescence,” Annu. Rev. Biophys. Bioeng. 13(1), 247–268 (1984).
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Supplementary Material (2)

NameDescription
» Visualization 1       TIRF of GFP-MreB in B. subtilis
» Visualization 2       TIRF-SIM of GFP-MreB in B. subtilis

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Figures (11)

Fig. 1.
Fig. 1. Modulation contrast maps CSTD(x,y) obtained via standard deviation of three grating images pm(x,y) at equidistant phases ϕm. (A) Measurement principle: two interfering laser beams (dark blue) generate the grating in object plane. From the images of the grating at equidistant phase positions the contrast map (B) is calculated, which is not corrected with a correction factor Acorr, yet. (C) Line profile of the TIRF-SIM gratings shown in (A). Averaging due to the large pixel size reduces the measured contrast.
Fig. 2.
Fig. 2. Scheme of dual color fast TIRF-SIM setup. The illumination unit includes illumination sources and beam steering devices (piezo scan mirror). The beam splitting unit comprises a Michelson interferometer with a retro reflector to mirror the beam at the optical axis. The polarization adjustment unit achieves the desired azimuthal polarization through a segmented polarizer (“Pizza polarizer”). In addition to the commercial microscope unit, there is custom-built detection unit, imaging two colors on separate sides of a sCMOS camera chip.
Fig. 3.
Fig. 3. Quantitative evaluation of the modulation contrast. (A) depicts possible configurations for contrast evaluation. Left: fluorescent probe. Right: illumination grating imaged directly via removal of emission filter (EF) and optional the dichroic mirror (DM). Optimal contrast is shown in (B) for dense fluorescent beads and in (D) for direct imaging of the illumination grating. (C) and (E) demonstrate the degradation of contrast due to a light source with insufficient spatial coherence, while (G) is an example for insufficient overlap of the two illumination beams due to misalignment. The last row visualizes the role of polarization and polarization dependent optics: (H) polarization 0° to incident plane of dichroic mirror. (I) Polarization turned by 90°. (J) dichroic mirror replaced by semi-transparent mirror for same polarization as in (I). Non-illuminated areas exhibit high standard deviations (dark red) due to camera noise.
Fig. 4.
Fig. 4. Imaging of bright fluorescent particles: resolution capabilities of TIRF-SIM. (A) and (D) show TIRF-SIM images of 92 nm fluorescent particles with an emission peak at 520 nm. Images (B), (E) and images (C), (F) show the corresponding TIRF and TIRF deconvolved images respectively. For comparison, (G) shows line scans of two neighbouring beads for the top and bottom row.
Fig. 5.
Fig. 5. Green fluorescent 92 nm beads imaged with different phase shifts between raw images of one illumination direction. For (A), (B) and (C) a phase shift of ${\textstyle{1 \over 3}}{\lambda _{ex}}_1 =$ 163 nm was used, whereas (E), (F) and (G) were acquired with a longer phase shift of ${\textstyle{1 \over 3}}{\bar{\lambda }_{sh}} =$ 173 nm between raw images of one direction. (B) and (F) are the corresponding image spectra and (C), (G) the average radial line scan of the spectra as indicated by the white lines in images of the spectra (B) and (F). (D) is the TIRF zero order image for comparison. Plot (H) depicts exemplary line scans for all three image modalities.
Fig. 6.
Fig. 6. Dual colour imaging of 200 nm beads. (A) and (B) show TIRF-SIM images taken simultaneously and sequentially respectively. (C) and (D) are TIRF (zero order) images for comparison, which are obtained from the nine raw images. The right column (C) and (E) are exemplary line scans for the green and red colour channel, respectively.
Fig. 7.
Fig. 7. Illumination contrast is reduced by sample movement. 85 nm fluorescent beads are moved at specific velocities during an acquisition of t0 = 100 ms for one raw image. (A) depicts the experimental scheme and (B) shows the theoretical contrast degradation of a moving fluorophore in an interference grating, while (C) depicts the actual intensity detected from the fluorophore, yielding an averaged grating and thus a lower contrast. (D) top row depicts the reconstructed images of the sample at various velocities. Bottom row shows TIRF zero order images for comparison. Below each image, the Fourier transform and a radial line scan is offered and the theoretical averaging of a single, infinitely small fluorophore moving across the grating. Scale bar is 1 µm.
Fig. 8.
Fig. 8. Similarity of TIRF-SIM images reconstructed from raw images with various signal-to-noise ratios (SNR). The SNR were obtained either experimentally by varying the illumination intensity or by computationally adding Gaussian noise to the raw images. Then, the images were cross-correlated, where the similarity parameter σ was obtained by extracting the peak value from the cross-correlation. Right: exemplary images p1(x,y) and p2(x,y).
Fig. 9.
Fig. 9. TIRF-SIM images of the same sample of raw images with different signal-to-noise ratios (SNR). The right box contains examples where SNR was experimentally varied by altering the illumination intensity. The left box shows TIRF-SIM images, which are reconstructed from raw data with added computational Gaussian noise. The upper and lower boxes compare two different Wiener filter parameters w used in the reconstruction process.
Fig. 10.
Fig. 10. MreB-Filaments in Bacillus subtilis spatially and temporally resolved with TIRF-SIM. (A) and (B) show the gain in spatial resolution, visualized by two exemplary line scans in (C). (D) depicts the dynamics of the three visible filaments. The filament indicated with a blue line is moving downwards and the two filaments marked with a green line are moving upwards. (E) and (F) show the kymographs along their tracks as a time representation. They reveal a unique stopping and reversal behaviour of the green filaments, while the blue filament constantly moves along its track.
Fig. 11.
Fig. 11. Long term, time series of TIRF-SIM images of (A) J774 macrophage cells with GFP life-Act labelling and (B) GPP labelled MreB filaments in Bacillus subtilis. Each time frame is colour coded visualizing the movement of actin filaments inside the cell (Scale bar is 0.5 µm – also see Visualization 1 and Visualization 2).

Equations (19)

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I e v , α ( r , z , ϕ 0 ) = ( I 1 ( r ) + I 2 ( r ) ) ( 1 + C m o d ( r ) cos ( 2   k e v , α r + ϕ 0 ) exp ( z / d ( θ i ) )
C m o d ( r ) = 2 I 1 ( r ) I 2 ( r ) I 0 ( r ) γ s ( r ) ,
I ( r , z ) = | E x ( r , z ) E y ( r , z ) E z ( r , z ) | 2 = | ( E 0 P ( r ) cos ( Ψ ) exp ( i ( δ p + π 2 ) ) f 1 ( θ i , n t i ) E 0 S ( r ) sin ( Ψ ) exp ( i δ s ) f 2 ( θ i , n t i ) E 0 P ( r ) cos ( Ψ ) exp ( i ( δ p + π 2 ) ) f 3 ( θ i , n t i ) ) | 2 exp ( z d ( θ i ) )
δ p = tan 1 ( sin 2 ( θ in ) n t i 2 n t i 2 cos ( θ in ) ) δ s = tan 1 ( sin 2 ( θ in ) n t i 2 cos ( θ in ) ) .
p ( x , y ) = ( L 0 ( x , y ) 0 d e z / d s ( x , y , z ) d z ) P S F ( x , y ) L 0 ( x , y ) s ( x , y , z = d 2 ) P S F ( x , y )
p m ( x , y ) σ f l I 0 , m ( x , y ) s ( x , y , z = d 2 ) P S F i n c ( x , y )
C S T D ( r ) = p S T D ( r ) p m e a n ( r )  =  1 M 1 m = 1 M ( p m ( r , ϕ m ) p m e a n ( r ) ) 2 p m e a n ( r ) .
F e v , s p h ( x , y , z , R ) = σ e x t Q f l R R R 2 x 2 R 2 x 2 R 2 x 2 y 2 R 2 x 2 y 2 I e v ( x , y , z ) c f l Q c o l ( z ) d x d y d z .
p m ( r , ϕ m ) = ( s ( r ) I e v ( r , ϕ m ) ) P S F ( r ) + n ( r ) .
P ~ m , α ( k , ϕ m ) = ( S ~ ( k ) I ~ e v , α ( k , ϕ m ) ) M T F ( k ) + N ~ ( k ) .
P ~ e f f ( k ) = α = 1 3 m = 1 1 S ~ ( k ± m 2 k e v , α ) e i ϕ m M T F ( k ) + N ~ ( k ) = S ~ ( k ) α = 1 3 m = 1 1 M T F ( k ± m 2 k e v , α ) + N ~ ( k ) = S ~ ( k ) M T F e f f ( k ) + N ~ ( k ) .
p S R ( r ) F T 1 [ S ~ ( k ) ] = F T 1 [ P ~ e f f ( k ) W ~ ( k ) A ~ ( k ) ]
W ~ ( k ) = M T F e f f ( k ) M T F e f f 2 ( k ) + w
S N R ( r ) = p S T D ( r ) p ¯ d a r k ( r ) p m e a n ( r ) .
C m o d ( x , y , Δ x , Δ y , Δ t ) = γ s ( Δ r , Δ t ) 2 E 1 ( x , y ) E 2 ( x + Δ x , y + Δ y ) / I 0 ( x , y )
I 0 , e v ( x , y , v x ) = 1 t 0 t 0 / 2 t 0 / 2 ( 1 + cos ( 2 k e v ( x v x t ) ) ) d t = s i n c ( k e v v x t 0 ) cos ( 2 k e v x ) + 1
v x 1 k e v t 0 = g π t 0
P e f f ( k , v x ) = | S ( k ) α = 1 1 m = 1 1 M T F ( k + m 2 k e v , α ) exp ( i m k e v v x t 0 ) |
P ~ e f f ( k , w ) = S ~ ( k ) A ~ ( k ) ( M T F e f f ( k ) + w M T F e f f ( k ) )