Enhanced 3D localization of individual RNA transcripts via astigmatic imaging

Evan P. Perillo, Leyma De Haro, Mary E. Phipps, Jennifer S. Martinez, Hsin Chih Yeh, Andrew K. Dunn, Douglas P. Shepherd, James H. Werner

Research output: Chapter in Book/Report/Conference proceedingConference contribution

1 Scopus citations

Abstract

Here we present an automated microscope capable of 3D multi-color single molecule localization of individual messenger RNA molecules in a wide range of cell types. We have implemented astigmatic imaging with a cylindrical lens to improve z-localization, and a maximum likelihood estimator on a graphics processing unit to improve localization precision and speed. This microscope will aid in gene expression analysis by its capability to perform high throughput imaging of thick cells and tissues while still maintaining sufficient z resolution to resolve single RNA transcripts in three dimensions. Enhanced z-localization allows for resolving membrane localized and co-localized transcripts.

Original languageEnglish (US)
Title of host publicationSingle Molecule Spectroscopy and Superresolution Imaging VII
PublisherSPIE
ISBN (Print)9780819498632
DOIs
StatePublished - 2014
Externally publishedYes
EventSingle Molecule Spectroscopy and Superresolution Imaging VII - San Francisco, CA, United States
Duration: Feb 1 2014Feb 2 2014

Publication series

NameProgress in Biomedical Optics and Imaging - Proceedings of SPIE
Volume8950
ISSN (Print)1605-7422

Conference

ConferenceSingle Molecule Spectroscopy and Superresolution Imaging VII
CountryUnited States
CitySan Francisco, CA
Period2/1/142/2/14

Keywords

  • Astigmatic imaging
  • Gene expression
  • Single-molecule localization
  • SmFISH
  • Uorescence microscopy

ASJC Scopus subject areas

  • Electronic, Optical and Magnetic Materials
  • Biomaterials
  • Atomic and Molecular Physics, and Optics
  • Radiology Nuclear Medicine and imaging

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