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Journal of Cell Science 114, 4153-4160 (2001)
© 2001 The Company of Biologists Limited

COMMENTARY

Cell biology beyond the diffraction limit: near-field scanning optical microscopy

Frank de Lange1, Alessandra Cambi1, Richard Huijbens1, Bärbel de Bakker2, Wouter Rensen2, Maria Garcia-Parajo2, Niek van Hulst2 and Carl G. Figdor1,*

1 Department of Tumor Immunology, University Medical Center Nijmegen, NCMLS/187 TIL, PO Box 9101, 6500HB Nijmegen, The Netherlands
2 Applied Optics Group, Faculty of Applied Physics and MESA+ Research Institute, University of Twente, PO Box 217, 7500AE Enschede, The Netherlands

*Author for correspondence (e-mail: c.figdor{at}mailbox.kun.nl)

Throughout the years, fluorescence microscopy has proven to be an extremely versatile tool for cell biologists to study live cells. Its high sensitivity and non-invasiveness, together with the ever-growing spectrum of sophisticated fluorescent indicators, ensure that it will continue to have a prominent role in the future. A drawback of light microscopy is the fundamental limit of the attainable spatial resolution – ~250 nm – dictated by the laws of diffraction. The challenge to break this diffraction limit has led to the development of several novel imaging techniques. One of them, near-field scanning optical microscopy (NSOM), allows fluorescence imaging at a resolution of only a few tens of nanometers and, because of the extremely small near-field excitation volume, reduces background fluorescence from the cytoplasm to the extent that single-molecule detection sensitivity becomes within reach. NSOM allows detection of individual fluorescent proteins as part of multimolecular complexes on the surface of fixed cells, and similar results should be achievable under physiological conditions in the near future.

Key words: Super-resolution, Near-field scanning optical microscopy, Single-molecule detection, Distribution, Cell surface, Membrane, GFP




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© The Company of Biologists Ltd 2001