English
 
Help Privacy Policy Disclaimer
  Advanced SearchBrowse

Item

ITEM ACTIONSEXPORT

Released

Journal Article

Flat-top TIRF illumination boosts DNA-PAINT imaging and quantification

MPS-Authors
/persons/resource/persons229551

Stehr,  Florian
Schwille, Petra / Cellular and Molecular Biophysics, Max Planck Institute of Biochemistry, Max Planck Society;

/persons/resource/persons229547

Stein,  Johannes
Schwille, Petra / Cellular and Molecular Biophysics, Max Planck Institute of Biochemistry, Max Planck Society;

/persons/resource/persons197583

Schueder,  Florian
Jungmann, Ralf / Molecular Imaging and Bionanotechnology, Max Planck Institute of Biochemistry, Max Planck Society;

/persons/resource/persons15815

Schwille,  Petra
Schwille, Petra / Cellular and Molecular Biophysics, Max Planck Institute of Biochemistry, Max Planck Society;

/persons/resource/persons172959

Jungmann,  Ralf
Jungmann, Ralf / Molecular Imaging and Bionanotechnology, Max Planck Institute of Biochemistry, Max Planck Society;

External Resource
No external resources are shared
Fulltext (restricted access)
There are currently no full texts shared for your IP range.
Fulltext (public)

s41467-019-09064-6.pdf
(Publisher version), 4MB

Supplementary Material (public)

41467_2019_9064_MOESM1_ESM.pdf
(Supplementary material), 4MB

Citation

Stehr, F., Stein, J., Schueder, F., Schwille, P., & Jungmann, R. (2019). Flat-top TIRF illumination boosts DNA-PAINT imaging and quantification. Nature Communications, 10: 1268. doi:10.1038/s41467-019-09064-6.


Cite as: http://hdl.handle.net/21.11116/0000-0003-DD12-0
Abstract
Super-resolution (SR) techniques have extended the optical resolution down to a few nanometers. However, quantitative treatment of SR data remains challenging due to its complex dependence on a manifold of experimental parameters. Among the different SR variants, DNA-PAINT is relatively straightforward to implement, since it achieves the necessary 'blinking' without the use of rather complex optical or chemical activation schemes. However, it still suffers from image and quantification artifacts caused by inhomogeneous optical excitation. Here we demonstrate that several experimental challenges can be alleviated by introducing a segment-wise analysis approach and ultimately overcome by implementing a flat-top illumination profile for TIRF microscopy using a commercially-available beam-shaping device. The improvements with regards to homogeneous spatial resolution and precise kinetic information over the whole field-of-view were quantitatively assayed using DNA origami and cell samples. Our findings open the door to high-throughput DNA-PAINT studies with thus far unprecedented accuracy for quantitative data interpretation.