Auto-Align - Multi-Modality Fluorescence Microscopy Image Co-registration

William T E Pitkeathly, Natalie S. Poulter, Ela Claridge, Joshua Z. Rappoport*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

5 Scopus citations


Multi-modality microscopes incorporate multiple microscopy techniques into one module, imaging through a common objective lens. Simultaneous or consecutive image acquisition of a single specimen, using multiple techniques, increases the amount of measurable information available. In order to benefit from each modality, it is necessary to accurately co-register data sets. Intrinsic differences in the image formation process employed by each modality result in images which possess different characteristics. In addition, as a result of using different measurement devices, images often differ in size and can suffer relative geometrical deformations including rotation, scale and translation, making registration a complex problem. Current methods generally rely on manual input and are therefore subject to human error. Here, we present an automated image registration tool for fluorescence microscopy. We show that it successfully registers images obtained via total internal reflection fluorescence (TIRF), or epi-fluorescence, and confocal microscopy. Furthermore, we provide several other applications including channel merging following image acquisition through an emission beam splitter, and lateral stage drift correction. We also discuss areas of membrane trafficking which could benefit from application of Auto-Align. Auto-Align is an essential item in the advanced microscopist's toolbox which can create a synergy of single or multi-modality image data.

Original languageEnglish (US)
Pages (from-to)204-217
Number of pages14
Issue number2
StatePublished - Feb 2012


  • Fluorescence microscopy
  • Image analysis
  • Image registration
  • Stage drift
  • Time lapse

ASJC Scopus subject areas

  • Structural Biology
  • Biochemistry
  • Molecular Biology
  • Genetics
  • Cell Biology

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