Molecular resolution imaging by post-labeling expansion single-molecule localization microscopy (Ex-SMLM)

Abstract

Expansion microscopy (ExM) enables super-resolution fluorescence imaging of physically expanded biological samples with conventional microscopes. By combining ExM with single-molecule localization microscopy (SMLM) it is potentially possible to approach the resolution of electron microscopy. However, current attempts to combine both methods remained challenging because of protein and fluorophore loss during digestion or denaturation, gelation, and the incompatibility of expanded polyelectrolyte hydrogels with photoswitching buffers. Here we show that re-embedding of expanded hydrogels enables d STORM imaging of expanded samples and demonstrate that post-labeling ExM resolves the current limitations of super-resolution microscopy. Using microtubules as a reference structure and centrioles, we demonstrate that post-labeling Ex-SMLM preserves ultrastructural details, improves the labeling efficiency and reduces the positional error arising from linking fluorophores into the gel thus paving the way for super-resolution imaging of immunolabeled endogenous proteins with true molecular resolution.

References 48

1. 1.
   Resolving bundled microtubules using anti-tubulin nanobodies · 2015

   No notice
2. 2.

   This reference’s wording can’t be shown under the paper’s licence.

   Not in our library

3. 3.

   This reference’s wording can’t be shown under the paper’s licence.

   Not in our library

4. 4.

   This reference’s wording can’t be shown under the paper’s licence.

   Not in our library

5. 5.

   This reference’s wording can’t be shown under the paper’s licence.

   Not in our library

6. 6.

   This reference’s wording can’t be shown under the paper’s licence.

   Not in our library

7. 7.

   This reference’s wording can’t be shown under the paper’s licence.

   Not in our library

8. 8.

   This reference’s wording can’t be shown under the paper’s licence.

   Not in our library

9. 9.

   This reference’s wording can’t be shown under the paper’s licence.

   Not in our library

10. 10.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

11. 11.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

12. 12.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

13. 13.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

14. 14.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

15. 15.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

16. 16.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

17. 17.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

18. 18.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

19. 19.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

20. 20.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

21. 21.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

22. 22.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

23. 23.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

24. 24.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

25. 25.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

26. 26.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

27. 27.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

28. 28.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

29. 29.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

30. 30.
    A peptide tag-specific nanobody enables high-quality labeling for dSTORM imaging · 2018

    No notice
31. 31.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

32. 32.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

33. 33.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

34. 34.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

35. 35.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

36. 36.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

37. 37.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

38. 38.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

39. 39.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

40. 40.

    This reference’s wording can’t be shown under the paper’s licence.

    Not in our library

Cited by papers we hold 6

• Magnify is a universal molecular anchoring strategy for expansion microscopy · 2023
  No notice
• iU-ExM: nanoscopy of organelles and tissues with iterative ultrastructure expansion microscopy · 2023
  No notice
• Analysis of super-resolution single molecule localization microscopy data: A tutorial · 2022
  Not checked
• Visualizing cellular and tissue ultrastructure using Ten-fold Robust Expansion Microscopy (TREx) · 2022
  Corrected
• Visualizing the native cellular organization by coupling cryofixation with expansion microscopy (Cryo-ExM) · 2022
  No notice
• Subdiffraction-resolution fluorescence imaging of immunological synapse formation between NK cells and A. fumigatus by expansion microscopy · 2021
  Not checked

Reads

A person counts once a day per paper. We keep no cookie and no address: a reader is a hash that changes every day. Any month under five reads shows no number.