Jul 08, 2025

Public workspaceLive imaging of primary neurons

Nature Communications
DOI
https://dx.doi.org/10.17504/protocols.io.261gek6eog47/v1
  • Bishal Basak1,2,
  • Erika Holzbaur1,2
  • 1Department of Physiology, Perelman School of Medicine, University of Pennsylvania, PA 19104;
  • 2Aligning Science Across Parkinson's (ASAP) Collaborative Research Network, Chevy Chase, MD 20815
  • University of Pennsylvania
Bishal Basak
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DOI: https://dx.doi.org/10.17504/protocols.io.261gek6eog47/v1
External link: https://doi.org/10.1038/s41467-025-62379-5
Protocol Citation: Bishal Basak, Erika Holzbaur 2025. Live imaging of primary neurons. protocols.io https://dx.doi.org/10.17504/protocols.io.261gek6eog47/v1
Manuscript citation:
Basak B, Holzbaur ELF (2025) Mitochondrial damage triggers the concerted degradation of negative regulators of neuronal autophagy. Nature Communications 16(). doi: 10.1038/s41467-025-62379-5
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: July 08, 2025
Last Modified: July 08, 2025
Protocol Integer ID: 221978
Keywords: live imaging of primary neurons live imaging, primary neurons live imaging, confocal microscope, primary neuron, live imaging, imaging
Funders Acknowledgements:
ASAP
Grant ID: ASAP-000350
Abstract
Live imaging of primary neurons under a confocal microscope
Troubleshooting
Imaging steps
Prior to imaging, warm 2 ml of imaging media per imaging dish at Temperature37 °C for atleast 15 minutes
(Imaging media composition:
45% glucose- 660 μl
B27- 1 ml
Hibernate E upto 50 ml)
During imaging, aspirate out conditioned media from the dishes, and add 2 ml of pre-warmed imaging media to the dish containing neurons
Image neurons live under a spinning disk confocal microscope at 100X magnification (oil objective)
Identify the soma of a neuron and collect z-stacks. Neurons which look unhealthy or show signs of blebbing should be excluded from imaging or further analysis.
Perform image analysis in Fiji/Image J based on experimental needs