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Localization accuracy in single molecule microscopy using electron-multiplying charge-coupled device cameras.

Jerry Chao1, E Sally Ward2, Raimund J Ober1

  • 1Dept. of Electrical Engineering, University of Texas at Dallas, Richardson, TX, USA ; Dept. of Immunology, University of Texas Southwestern Medical Center, Dallas, TX, USA.

Proceedings of Spie--The International Society for Optical Engineering
|January 1, 2014
PubMed
Summary
This summary is machine-generated.

We developed a theory to calculate the accuracy limit for estimating parameters from electron-multiplying charge-coupled device (EMCCD) images. This provides a method to compare detectors like EMCCD and charge-coupled device (CCD) for low light imaging.

Keywords:
Branching processCramer-Rao lower boundFisher information matrixelectron multiplicationsingle molecule microscopy

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Area of Science:

  • Scientific Imaging
  • Optical Microscopy
  • Statistical Optics

Background:

  • Electron-multiplying charge-coupled devices (EMCCDs) are crucial for low light imaging, particularly in single molecule microscopy.
  • Estimating parameters (e.g., molecular location) from EMCCD images has lacked a theoretical framework for accuracy assessment.
  • Conventional charge-coupled devices (CCDs) are limited by readout noise in low light conditions.

Purpose of the Study:

  • To develop a theoretical framework for calculating the Cramer-Rao lower bound on parameter estimation accuracy from EMCCD images.
  • To introduce a "noise coefficient" for comparing the Fisher information of different detector models.
  • To assess the localization accuracy of single molecules using EMCCD imaging.

Main Methods:

  • Developed theory for Fisher information matrix calculation in EMCCD imaging.
  • Modeled the stochastic electron multiplication process as a geometrically multiplied branching process.
  • Applied the theory to single molecule localization and compared theoretical limits with simulation results.

Main Results:

  • Established a method to determine the theoretical accuracy limit for parameter estimation in EMCCD images.
  • Introduced a "noise coefficient" to objectively compare EMCCD and CCD detector performance across various signal levels.
  • Calculated localization accuracy limits for single molecules, validated against maximum likelihood estimates from simulated data.

Conclusions:

  • The developed theory provides a robust method for evaluating parameter estimation accuracy in EMCCD imaging.
  • The "noise coefficient" offers a universal metric for selecting optimal detectors (EMCCD vs. CCD) based on signal conditions.
  • This work advances the understanding of EMCCD capabilities for high-precision measurements in low light microscopy.