Method for co-cluster analysis in multichannel single-molecule localisation data

Histochem Cell Biol. 2014 Jun;141(6):605-12. doi: 10.1007/s00418-014-1208-z. Epub 2014 Mar 19.

Abstract

We demonstrate a combined univariate and bivariate Getis and Franklin's local point pattern analysis method to investigate the co-clustering of membrane proteins in two-dimensional single-molecule localisation data. This method assesses the degree of clustering of each molecule relative to its own species and relative to a second species. Using simulated data, we show that this approach can quantify the degree of cluster overlap in multichannel point patterns. The method is validated using photo-activated localisation microscopy and direct stochastic optical reconstruction microscopy data of the proteins Lck and CD45 at the T cell immunological synapse. Analysing co-clustering in this manner is generalizable to higher numbers of fluorescent species and to three-dimensional or live cell data sets.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Humans
  • Image Processing, Computer-Assisted
  • Immunological Synapses / metabolism*
  • Jurkat Cells
  • Leukocyte Common Antigens / analysis
  • Leukocyte Common Antigens / metabolism
  • Lymphocyte Specific Protein Tyrosine Kinase p56(lck) / analysis
  • Lymphocyte Specific Protein Tyrosine Kinase p56(lck) / metabolism
  • Microscopy, Fluorescence / methods*
  • T-Lymphocytes / cytology
  • T-Lymphocytes / metabolism

Substances

  • Lymphocyte Specific Protein Tyrosine Kinase p56(lck)
  • Leukocyte Common Antigens