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Determining Cell Number During Cell Culture using the Scepter Cell Counter
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Cell colony counter called CoCoNut.

Mattia Siragusa1, Stefano Dall'Olio2, Pil M Fredericia1

  • 1The Hevesy Laboratory, Center for Nuclear Technologies, Technical University of Denmark, Roskilde, Denmark.

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|November 8, 2018
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Summary

This study introduces CoCoNut, a semi-automated system for counting cell colonies, significantly speeding up analysis after radiation damage. CoCoNut offers a user-friendly, cost-effective solution for reproducible clonogenic assay results.

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Area of Science:

  • Cell Biology
  • Radiation Biology
  • Bio-imaging

Background:

  • Clonogenic assays are vital for assessing cell reproductive death post-biological damage, particularly from ionizing radiation.
  • Traditional manual colony counting is time-consuming, subjective, and visually demanding.
  • Existing automated methods may lack accuracy or require complex calibration.

Purpose of the Study:

  • To develop and validate a semi-automated, image-based system (CoCoNut) for efficient and objective cell colony counting.
  • To improve the speed and reproducibility of clonogenic assay analysis.
  • To provide a cost-effective and user-friendly alternative to manual counting.

Main Methods:

  • CoCoNut utilizes an ImageJ macro combined with a 3D-printed light-box for Crystal Violet-stained colonies.
  • Image acquisition employs optimal lighting and a novel background removal technique (dividing colony image by empty flask image).
  • The system requires minimal calibration, adjusting only the smallest colony area parameter.

Main Results:

  • CoCoNut accurately distinguishes between single and merged colonies and identifies edge colonies.
  • The semi-automated system demonstrated robust performance with V79 and HeLa cell samples.
  • Results showed high reproducibility and comparability across different settings, outperforming other freely available tools.

Conclusions:

  • CoCoNut offers a fast, inexpensive, and user-friendly solution for semi-automated cell colony counting.
  • The system enhances objectivity and reproducibility in clonogenic assays.
  • The combination of software and a reproducible 3D-printed light-box makes CoCoNut an accessible tool for research.