Imperial College London

Professor Nick Heard

Faculty of Natural SciencesDepartment of Mathematics

Chair in Statistics



+44 (0)20 7594 1490n.heard Website




543Huxley BuildingSouth Kensington Campus






BibTex format

author = {Griffié, J and Shannon, M and Bromley, CL and Boelen, L and Burn, GL and Williamson, DJ and Heard, NA and Cope, AP and Owen, DM and Rubin-Delanchy, P},
doi = {10.1038/nprot.2016.149},
journal = {Nature Protocols},
pages = {2499--2514},
title = {A Bayesian cluster analysis method for single-molecule localization microscopy data},
url = {},
volume = {11},
year = {2016}

RIS format (EndNote, RefMan)

AB - Cell function is regulated by the spatiotemporal organization of the signaling machinery, and a key facet of this is molecular clustering. Here, we present a protocol for the analysis of clustering in data generated by 2D single-molecule localization microscopy (SMLM)-for example, photoactivated localization microscopy (PALM) or stochastic optical reconstruction microscopy (STORM). Three features of such data can cause standard cluster analysis approaches to be ineffective: (i) the data take the form of a list of points rather than a pixel array; (ii) there is a non-negligible unclustered background density of points that must be accounted for; and (iii) each localization has an associated uncertainty in regard to its position. These issues are overcome using a Bayesian, model-based approach. Many possible cluster configurations are proposed and scored against a generative model, which assumes Gaussian clusters overlaid on a completely spatially random (CSR) background, before every point is scrambled by its localization precision. We present the process of generating simulated and experimental data that are suitable to our algorithm, the analysis itself, and the extraction and interpretation of key cluster descriptors such as the number of clusters, cluster radii and the number of localizations per cluster. Variations in these descriptors can be interpreted as arising from changes in the organization of the cellular nanoarchitecture. The protocol requires no specific programming ability, and the processing time for one data set, typically containing 30 regions of interest, is ∼18 h; user input takes ∼1 h.
AU - Griffié,J
AU - Shannon,M
AU - Bromley,CL
AU - Boelen,L
AU - Burn,GL
AU - Williamson,DJ
AU - Heard,NA
AU - Cope,AP
AU - Owen,DM
AU - Rubin-Delanchy,P
DO - 10.1038/nprot.2016.149
EP - 2514
PY - 2016///
SN - 1754-2189
SP - 2499
TI - A Bayesian cluster analysis method for single-molecule localization microscopy data
T2 - Nature Protocols
UR -
UR -
UR -
VL - 11
ER -