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A variable, usually notated by capital letters such as X and Y, is a characteristic or measurement that can be determined for each member of a population. Data are the actual values of variables. They may be numbers, or they may be words. Datum is a single value.
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SPEKcheck - fluorescence microscopy spectral visualisation and optimisation: a web application, javascript library,

Mick A Phillips1, David M Susano Pinto1, Ian M Dobbie1

  • 1Micron Advanced Bioimaging Unit, Department of Biochemistry, University of Oxford, South Parks Road, Oxford, OX13QU, UK.

Wellcome Open Research
|September 4, 2018
PubMed
Summary
This summary is machine-generated.

SPEKcheck is a new web application that helps researchers optimize fluorescence microscopy by visualizing light path efficiency. It aids in selecting optimal dye and filter combinations for better multicolour imaging.

Keywords:
ExcitationFluorescenceMicroscopyModelling

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Area of Science:

  • Biophysics
  • Microscopy
  • Optical Engineering

Background:

  • Advanced fluorescence microscopy demands precise alignment of optical components for optimal performance.
  • Selecting appropriate filter sets and fluorophores is critical, especially for complex multicolour imaging, but often relies on empirical methods.

Purpose of the Study:

  • To develop a user-friendly web application, SPEKcheck, for evaluating fluorescence microscope light path efficiency.
  • To provide a tool that aids in selecting optimal dye-filter combinations and troubleshooting spectral overlap issues.

Main Methods:

  • Developed SPEKcheck, a web-based application for visualizing fluorescence microscope light path efficiency.
  • The software calculates excitation efficiency, emission collection efficiency, and a composite "brightness" score for various fluorophores.
  • It generates spectral plots to identify potential issues like low excitation/emission efficiency and high bleedthrough.

Main Results:

  • SPEKcheck provides quantitative metrics for comparing the performance of different fluorescent labels and filter sets.
  • The application visualizes spectral data, enabling users to easily identify and mitigate potential problems in multicolour fluorescence imaging.
  • It facilitates informed decision-making for optimizing microscope configurations.

Conclusions:

  • SPEKcheck offers a valuable resource for researchers to optimize fluorescence microscope performance.
  • The tool simplifies the selection of optimal dye-filter combinations, improving the efficiency and reliability of multicolour fluorescence imaging.
  • It addresses a critical need for systematic evaluation of optical configurations in fluorescence microscopy.