title: Local dimensionality determines imaging speed in localization microscopy.
abstract: Localization microscopy allows biological samples to be imaged at a length scale of tens of nanometres. Live-cell super-resolution imaging is rare, as it is generally assumed to be too slow for dynamic samples. The speed of data acquisition can be optimized by tuning the density of activated fluorophores in each time frame. Here, we show that the maximum achievable imaging speed for a particular structure varies by orders of magnitude, depending on the sample dimensionality (that is, whether the sample is more like a point, a strand or an extended structure such as a focal adhesion). If too high an excitation density is used, we demonstrate that the analysis undergoes silent failure, resulting in reconstruction artefacts. We are releasing a tool to allow users to identify areas of the image in which the activation density was too high and correct for them, in both live- and fixed-cell experiments.
authors.affiliation: Randall Division, King's College London, New Hunts House, Guy's Campus, London SE1 1UL, UK.
authors.forename: Patrick
authors.initials: P
authors.lastname: Fox-Roberts
authors.affiliation: Randall Division, King's College London, New Hunts House, Guy's Campus, London SE1 1UL, UK.
authors.forename: Richard
authors.initials: R
authors.lastname: Marsh
authors.affiliation: Randall Division, King's College London, New Hunts House, Guy's Campus, London SE1 1UL, UK.
authors.forename: Karin
authors.initials: K
authors.lastname: Pfisterer
authors.affiliation: Randall Division, King's College London, New Hunts House, Guy's Campus, London SE1 1UL, UK.
authors.forename: Asier
authors.identifier: 0000-0002-9899-8723
authors.initials: A
authors.lastname: Jayo
authors.affiliation: Randall Division, King's College London, New Hunts House, Guy's Campus, London SE1 1UL, UK.
authors.forename: Maddy
authors.initials: M
authors.lastname: Parsons
authors.affiliation: Randall Division, King's College London, New Hunts House, Guy's Campus, London SE1 1UL, UK.
authors.forename: Susan
authors.initials: S
authors.lastname: Cox
mesh_terms: D016477:Artifacts; D003198:Computer Simulation; D006367:HeLa Cells; D006801:Humans; D008856:Microscopy, Fluorescence / Q000379:methods*; D008962:Models, Theoretical