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Varying label density allows artifact-free analysis of membrane-protein nanoclusters.

ABSTRACT: We present a method to robustly discriminate clustered from randomly distributed molecules detected with techniques based on single-molecule localization microscopy, such as PALM and STORM. The approach is based on deliberate variation of labeling density, such as titration of fluorescent antibody, combined with quantitative cluster analysis, and it thereby circumvents the problem of cluster artifacts generated by overcounting of blinking fluorophores. The method was used to analyze nanocluster formation in resting and activated immune cells.

SUBMITTER: Baumgart F

PROVIDER: S-EPMC6404959 | biostudies-literature | 2016 Aug

REPOSITORIES: biostudies-literature

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Varying label density allows artifact-free analysis of membrane-protein nanoclusters.

Baumgart Florian F Arnold Andreas M AM Leskovar Konrad K Staszek Kaj K Fölser Martin M Weghuber Julian J Stockinger Hannes H Schütz Gerhard J GJ

Nature methods 20160613 8

We present a method to robustly discriminate clustered from randomly distributed molecules detected with techniques based on single-molecule localization microscopy, such as PALM and STORM. The approach is based on deliberate variation of labeling density, such as titration of fluorescent antibody, combined with quantitative cluster analysis, and it thereby circumvents the problem of cluster artifacts generated by overcounting of blinking fluorophores. The method was used to analyze nanocluster ...[more]

PMID: 27295310

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