title: An information-theoretic approach to designing the plane spacing for multifocal plane microscopy.
abstract: Multifocal plane microscopy (MUM) is a 3D imaging modality which enables the localization and tracking of single molecules at high spatial and temporal resolution by simultaneously imaging distinct focal planes within the sample. MUM overcomes the depth discrimination problem of conventional microscopy and allows high accuracy localization of a single molecule in 3D along the z-axis. An important question in the design of MUM experiments concerns the appropriate number of focal planes and their spacings to achieve the best possible 3D localization accuracy along the z-axis. Ideally, it is desired to obtain a 3D localization accuracy that is uniform over a large depth and has small numerical values, which guarantee that the single molecule is continuously detectable. Here, we address this concern by developing a plane spacing design strategy based on the Fisher information. In particular, we analyze the Fisher information matrix for the 3D localization problem along the z-axis and propose spacing scenarios termed the strong coupling and the weak coupling spacings, which provide appropriate 3D localization accuracies. Using these spacing scenarios, we investigate the detectability of the single molecule along the z-axis and study the effect of changing the number of focal planes on the 3D localization accuracy. We further review a software module we recently introduced, the MUMDesignTool, that helps to design the plane spacings for a MUM setup.
authors.affiliation: Dept. of Biomedical Engineering, Texas A&M University, College Station, TX, USA ; Dept. of Molecular and Cellular Medicine, Texas A&M Health Science Center, College Station, TX, USA.
authors.forename: Amir
authors.initials: A
authors.lastname: Tahmasbi
authors.affiliation: Dept. of Immunology, University of Texas Southwestern Medical Center, Dallas, TX, USA.
authors.forename: Sripad
authors.initials: S
authors.lastname: Ram
authors.affiliation: Dept. of Biomedical Engineering, Texas A&M University, College Station, TX, USA ; Dept. of Molecular and Cellular Medicine, Texas A&M Health Science Center, College Station, TX, USA.
authors.forename: Jerry
authors.initials: J
authors.lastname: Chao
authors.affiliation: Dept. of Biomedical Engineering, Texas A&M University, College Station, TX, USA ; Dept. of Molecular and Cellular Medicine, Texas A&M Health Science Center, College Station, TX, USA.
authors.forename: Anish V
authors.initials: AV
authors.lastname: Abraham
authors.affiliation: Dept. of Molecular and Cellular Medicine, Texas A&M Health Science Center, College Station, TX, USA ; Dept. of Microbial Pathogenesis and Immunology, Texas A&M Health Science Center, College Station, TX, USA.
authors.forename: E Sally
authors.initials: ES
authors.lastname: Ward
authors.affiliation: Dept. of Biomedical Engineering, Texas A&M University, College Station, TX, USA ; Dept. of Molecular and Cellular Medicine, Texas A&M Health Science Center, College Station, TX, USA.
authors.forename: Raimund J
authors.initials: RJ
authors.lastname: Ober
keywords: Cramér-Rao lower bound; Fisher Information Matrix; Single molecule microscopy