Direct Supercritical Angle Localization Microscopy for Nanometer 3D Superresolution

Dasgupta, Anindita; Deschamps, Joran; Matti, Ulf; Huebner, Uwe; Becker, J.; Strauss, Sebastian; Jungmann, Ralf; Heintzmann, Rainer; Ries, Jonas

doi:10.1101/2020.06.25.171058

Cited by 2 publications

(4 citation statements)

References 40 publications

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“…near-field emission that is converted to propagating light by interacting with the boundary) to further enhance their resolution calculation (i.e. the detection NA is limited to the refractive index of the sample), despite other measurements with the same objective showing that significant super-critical light can be present [40].…”

Section: Should High-na Tirf-sim Which Can Achieve Lateral Resolution...mentioning

confidence: 99%

Answering some questions about structured illumination microscopy

Manton

2022

Phil. Trans. R. Soc. A.

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Structured illumination microscopy (SIM) provides images of fluorescent objects at an enhanced resolution greater than that of conventional epifluorescence wide-field microscopy. Initially demonstrated in 1999 to enhance the lateral resolution twofold, it has since been extended to enhance axial resolution twofold (2008), applied to live-cell imaging (2009) and combined with myriad other techniques, including interferometric detection (2008), confocal microscopy (2010) and light sheet illumination (2012). Despite these impressive developments, SIM remains, perhaps, the most poorly understood ‘super-resolution’ method. In this article, we provide answers to the 13 questions regarding SIM proposed by Prakash et al. along with answers to a further three questions. After providing a general overview of the technique and its developments, we explain why SIM as normally used is still diffraction-limited. We then highlight the necessity for a non-polynomial, and not just nonlinear, response to the illuminating light in order to make SIM a true, diffraction-unlimited, super-resolution technique. In addition, we present a derivation of a real-space SIM reconstruction approach that can be used to process conventional SIM and image scanning microscopy (ISM) data and extended to process data with quasi-arbitrary illumination patterns. Finally, we provide a simple bibliometric analysis of SIM development over the past two decades and provide a short outlook on potential future work. This article is part of the Theo Murphy meeting issue ‘Super-resolution structured illumination microscopy (part 2)’.

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Section: Should High-na Tirf-sim Which Can Achieve Lateral Resolution...mentioning

confidence: 99%

Answering some questions about structured illumination microscopy

Manton

2022

Phil. Trans. R. Soc. A.

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“…Referring the calculations to a constant UAF signal rather to a constant total signal (i.e. UAF + SAF) provides more realistic curves for fluorophores with low quantum yield, where UAF and SAF signals are not competing [ 32 ]. For high quantum yields, the onset of SAF will somewhat decrease UAF, but the resulting changes to the CRLB curves are be minor.…”

Section: Localization Performance Close To the Coverslipmentioning

confidence: 99%

“…In addition, we have performed simulations on the only recently experimentally demonstrated method dSALM, where SAF and UAF contributions are separately imaged without blocking any light [ 32 ]. Since the performance of dSALM depends strongly on the background levels of the two imaging channels and thus also on the illumination conditions (TIRF vs. epi-fluorescence excitation at low angles), we refrain from including the method in Fig.…”

Section: Localization Performance Close To the Coverslipmentioning

confidence: 99%

“…Other methods known as SALM [ 30 ], respectively DONALD [ 31 ] or the recently introduced direct SALM (dSALM) method [ 32 ] draw information from supercritical angle fluorescence (SAF), i.e., the part of the non-propagating near field turning into propagating waves inside the coverslip whenever an emitter is sufficiently close. To measure SAF, SALM and dSALM employ beam splitting and two-channel detection.…”

Section: Introductionmentioning

confidence: 99%

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Three-dimensional single molecule localization close to the coverslip: a comparison of methods exploiting supercritical angle fluorescence

Zelger¹,

Bodner²,

Offterdinger³

et al. 2021

Biomed. Opt. Express

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The precise spatial localization of single molecules in three dimensions is an important basis for single molecule localization microscopy (SMLM) and tracking. At distances up to a few hundred nanometers from the coverslip, evanescent wave coupling into the glass, also known as supercritical angle fluorescence (SAF), can strongly improve the axial precision, thus facilitating almost isotropic localization performance. Specific detection systems, introduced as Supercritical angle localization microscopy (SALM) or Direct optical nanoscopy with axially localized detection (DONALD), have been developed to exploit SAF in modified two-channel imaging schemes. Recently, our group has shown that off-focus microscopy, i.e., imaging at an intentional slight defocus, can perform equally well, but uses only a single detection arm. Here we compare SALM, off-focus imaging and the most commonly used 3D SMLM techniques, namely cylindrical lens and biplane imaging, regarding 3D localization in close proximity to the coverslip. We show that all methods gain from SAF, which leaves a high detection NA as the only major key requirement to unlock the SAF benefit. We find parameter settings for cylindrical lens and biplane imaging for highest z-precision. Further, we compare the methods in view of robustness to aberrations, fixed dipole emission and double-emitter events. We show that biplane imaging provides the best overall performance and support our findings by DNA-PAINT experiments on DNA-nanoruler samples. Our study sheds light on the effects of SAF for SMLM and is helpful for researchers who plan to employ localization-based 3D nanoscopy close to the coverslip.

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Direct Supercritical Angle Localization Microscopy for Nanometer 3D Superresolution

Cited by 2 publications

References 40 publications

Answering some questions about structured illumination microscopy

Answering some questions about structured illumination microscopy

Three-dimensional single molecule localization close to the coverslip: a comparison of methods exploiting supercritical angle fluorescence

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