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Live-Cell Imaging: A Balancing Act Between Speed, Sensitivity, and Resolution

What You Will Learn in This Chapter

Live-cell imaging is perhaps one of the most exciting and challenging activities in the field of microscopy. It is exciting as recent developments in microscope technology have enabled scientists to visualize cellular and subcellular processes in real time down to the molecular level. With this comes the prospect of studying the mechanisms of diseases in greater detail and finding possible therapeutic solutions. Nevertheless, live-cell imaging is equally challenging because cells themselves and in fact—all cellular processes—are extremely sensitive to the very impact of using light for their visualization. The aim of this chapter is to provide a practical overview for early PhD students as well as more experienced post-docs, who will spend considerable time mastering the most important challenges and prerequisites in the very rapidly evolving field of live-cell microscopy.


  • Live cells
  • Microscopy
  • Phototoxicity
  • Controlled environment
  • Focus drift
  • Fluorescent probes
  • Automation

“In honour of Professor Anirban Banerjee, who taught us the wonders of observing living cells…”

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Change history

  • 29 January 2023

    The original version of this chapter was inadvertently published with errors. The following corrections have been made after publication.



Acousto-optical tunable filter




Charge-coupled device


Chromosome instability


Complementary metal-oxide-semiconductor


Confocal microscopy


Colorectal cancer


Clustered regularly interspaced short palindromic repeats


CRISPR-Cas9-mediated homology-independent organoid transgenesis


Confocal scanning laser microscopy




Differential interference contrast


Electron multiplying charged-coupled device


Epifluorescence microscopy




Field of view


Fluorescence recovery after photobleaching


Fluorescence resonance energy transfer


Green fluorescent protein


High-content analysis


High-content screening


High-efficiency particulate air


Human umbilical vein endothelial cells




Light-emitting diode


Numerical aperture




Perfect focus system


Photomultiplier tube


Point spread function


Pumilio-homology domain


Polydactyl zinc finger


Re-scanning confocal microscopy


Red fluorescent protein


Region of interest


Structured illumination microscopy


Signal-to-noise ratio


Stimulated emission depletion microscopy


Stochastic optical reconstruction microscopy


Transcription activator-like effector


Total internal reflection fluorescence


Transistor-transistor logic




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Appendix: Microscope Company and Resources List with Internet-Links

Appendix: Microscope Company and Resources List with Internet-Links

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Kole, J., Ahmed, H., Chatterjee, N., Lukinavičius, G., Musters, R. (2022). Live-Cell Imaging: A Balancing Act Between Speed, Sensitivity, and Resolution. In: Nechyporuk-Zloy, V. (eds) Principles of Light Microscopy: From Basic to Advanced . Springer, Cham.

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