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Multiplexed Lectin-PAINT super-resolution microscopy enables cell glycotyping.

Marrit M E Tholen1, Roger Riera1, Cristina Izquierdo-Lozano1

  • 1Department of Biomedical Engineering, Institute for Complex Molecular Systems (ICMS), Eindhoven University of Technology, Eindhoven, 5612AZ, The Netherlands.

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We developed Lectin-PAINT, a super-resolution imaging technique for visualizing cell surface sugars (glycans) in live cells. This method allows detailed mapping of cellular glycans, revealing differences between cancer cell types.

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Area of Science:

  • Cellular Biology
  • Glycobiology
  • Super-resolution Microscopy

Background:

  • Glycosylation is crucial for cellular functions, but its complex patterns are difficult to study.
  • Existing methods lack sensitivity, multiplexing, or in-situ single-cell analysis capabilities for glycans.

Purpose of the Study:

  • To introduce Lectin-PAINT, a novel super-resolution imaging method for live-cell visualization of the cellular glycocalyx.
  • To enable multiplexed, single-cell, and single-molecule level analysis of cell surface carbohydrates.

Main Methods:

  • Lectin-PAINT utilizes the reversible binding of lectins to specific carbohydrate structures for super-resolution imaging.
  • Employs a tailored lectin library for multiplexed detection of diverse carbohydrate families.
  • Combines 8-color super-resolution imaging with Point Accumulation In Nanoscale Topography (PAINT) principles.

Main Results:

  • Achieved super-resolution imaging of the glycocalyx at the single-molecule level in live cells.
  • Quantified over 350 glycosylation parameters per cell, defining a unique 'glycotype' or glycan fingerprint.
  • Demonstrated glytyping of various cancer cell types, revealing significant heterogeneity and variability in their glycocalyx.

Conclusions:

  • Lectin-PAINT overcomes limitations of previous techniques, offering unprecedented detail in glycocalyx analysis.
  • The 'glycotype' provides a powerful tool for understanding cell surface glycan heterogeneity in biological contexts, particularly in cancer.
  • This research lays the foundation for understanding disease-associated changes in the glycocalyx.