Automatic fluorescent tag localization II: Improvement in super-resolution by relative tracking

D. Thomann, J. Dorn, P. K. Sorger, Gaudenz Danuser

Research output: Contribution to journalArticle

45 Citations (Scopus)

Abstract

We present an algorithm for the three-dimensional (3D) tracking of multiple fluorescent subresolution tags with super-resolution in images of living cells. Recently, we described an algorithm for the automatic detection of such tags in single flames and demonstrated its potential in a biological system. The algorithm presented here adds to the tag detector a module for relative tracking of the signals between frames. As with tag detection, the main problem in relative tracking arises when signals of multiple tags interfere. We propose a novel multitemplate matching framework that exploits knowledge of the microscope point spread function to separate the intensity contribution of each tag in image regions with signal interferences. We use this intensity splitting to reconstruct a template for each tag in the source frame and a patch in the target frame, which are both free of intensity contributions from other tag signals. Tag movements between flames are then tracked by seeking, for each template-patch pair, the displacement vector providing the best signal match in terms of the sum of squared intensity differences. Because template and patch generation of tags with overlapping signals are interdependent, the matching is carried out simultaneously for all tags, and in an iterative manner. We have examined the performance of our approach using synthetic 3D data and observed a significant increase in resolution and robustness as compared with our previously described detector. It is now possible to localize and track tags separated by a distance three times smaller than the Rayleigh limit with a relative positional accuracy of better than 50 nm. We have applied the new tracking system to extract metaphase trajectories of fluorescently tagged chromosomes relative to the spindle poles in budding yeast.

Original languageEnglish (US)
Pages (from-to)230-248
Number of pages19
JournalJournal of Microscopy
Volume211
Issue number3
StatePublished - Sep 1 2003

Fingerprint

Spindle Poles
Detectors
Saccharomycetales
templates
Optical transfer function
Biological systems
Signal interference
Metaphase
Chromosomes
Yeast
flames
Poles
Microscopes
Cells
Trajectories
spindles
chromosomes
detectors
yeast
point spread functions

Keywords

  • Computer vision
  • GFP tags
  • Super-resolution
  • Tracking

ASJC Scopus subject areas

  • Instrumentation

Cite this

Automatic fluorescent tag localization II : Improvement in super-resolution by relative tracking. / Thomann, D.; Dorn, J.; Sorger, P. K.; Danuser, Gaudenz.

In: Journal of Microscopy, Vol. 211, No. 3, 01.09.2003, p. 230-248.

Research output: Contribution to journalArticle

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