Automated 5-D analysis of cell migration and interaction in the thymic cortex from time-lapse sequences of 3-D multi-channel multi-photon images

J Immunol Methods. 2009 Jan 1;340(1):65-80. doi: 10.1016/j.jim.2008.09.024. Epub 2008 Nov 4.

Abstract

This paper presents automated methods to quantify dynamic phenomena such as cell-cell interactions and cell migration patterns from time-lapse series of multi-channel three-dimensional image stacks of living specimens. Various 5-dimensional (x, y, z, t, lambda) images containing dendritic cells (DC), and T-cells or thymocytes in the developing mouse thymic cortex and lymph node were acquired by two-photon laser scanning microscopy (TPLSM). The cells were delineated automatically using a mean-shift clustering algorithm. This enables morphological measurements to be computed. A robust multiple-hypothesis tracking algorithm was used to track thymocytes (the DC were stationary). The tracking data enable dynamic measurements to be computed, including migratory patterns of thymocytes, and duration of thymocyte-DC contacts. Software was developed for efficient inspection, corrective editing, and validation of the automated analysis results. Our software-generated results agreed with manually generated measurements to within 8%.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Algorithms
  • Animals
  • Cell Movement / immunology*
  • Chimera
  • Dendritic Cells / cytology
  • Dendritic Cells / immunology
  • Image Processing, Computer-Assisted / methods
  • Mice
  • Mice, Inbred C57BL
  • Mice, Inbred DBA
  • Mice, Transgenic
  • Microscopy, Confocal / methods
  • Microscopy, Fluorescence, Multiphoton / methods
  • T-Lymphocytes / cytology
  • T-Lymphocytes / immunology
  • Thymus Gland / cytology*
  • Thymus Gland / immunology*