Simultaneous imaging of different focal planes in fluorescence microscopy for the study of cellular dynamics in three dimensions

IEEE Trans Nanobioscience. 2004 Dec;3(4):237-42. doi: 10.1109/tnb.2004.837899.


The imaging of cellular dynamics in three dimensions using a standard microscope is severely limited due to the fact that only one focal plane can be imaged at a given point in time. Here we present a modification of the classical microscope design with which two or more focal planes can be imaged simultaneously. This is achieved by a modification of the emission pathway of a standard microscope. The efficacy of the design is shown by imaging bead samples and an FcRn-green fluorescent protein expressing tubule that leaves a sorting endosome and subsequently exocytoses at the plasma membrane.

Publication types

  • Evaluation Study
  • Research Support, U.S. Gov't, P.H.S.
  • Validation Study

MeSH terms

  • Cell Line
  • Endothelial Cells / cytology*
  • Endothelial Cells / metabolism*
  • Equipment Design
  • Equipment Failure Analysis
  • Histocompatibility Antigens Class I
  • Humans
  • Image Enhancement / instrumentation*
  • Imaging, Three-Dimensional / instrumentation*
  • Imaging, Three-Dimensional / methods
  • Microscopy, Fluorescence / instrumentation*
  • Microscopy, Fluorescence / methods
  • Microscopy, Video / instrumentation*
  • Microscopy, Video / methods
  • Receptors, Fc / metabolism*
  • Reproducibility of Results
  • Sensitivity and Specificity


  • Histocompatibility Antigens Class I
  • Receptors, Fc
  • Fc receptor, neonatal