Feb 26, 2026

Public workspaceSynchronization Protocol - Eggert lab

DOI
https://dx.doi.org/10.17504/protocols.io.rm7vze21rvx1/v1
  • Andrea Paquola1,2,
  • Ulrike Eggert1,2
  • 1Randall Centre for Cell and Molecular Biophysics, King’s College London, New Hunt’s House, Guy’s Campus, SE1 1UL, UK;
  • 2Department of Chemistry, King’s College London, London, UK, SE1 1UL, UK
  • Andrea Paquola
Andrea Paquola
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DOI: https://dx.doi.org/10.17504/protocols.io.rm7vze21rvx1/v1
Protocol Citation: Andrea Paquola, Ulrike Eggert 2026. Synchronization Protocol - Eggert lab. protocols.io https://dx.doi.org/10.17504/protocols.io.rm7vze21rvx1/v1
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: February 17, 2026
Last Modified: February 26, 2026
Protocol Integer ID: 243402
Keywords: synchronization of hela cell, eggert lab this protocol, hela cell, synchronization protocol, using nocodazole, synchronization, cytokinesi, eggert lab, protocol
Funders Acknowledgements:
Wellcome Trust
Grant ID: 110060/Z/15/Z
BBSRC
Grant ID: BB/V003518/1
Abstract
This protocol describes the synchronization of HeLa cells at cytokinesis using nocodazole.
Troubleshooting
Synchronization
Plate HeLa cells at a density of 4x106 cells per T75 flask.
The following day, add 100 ng/mL nocodazole (Sigma-Aldrich) to 20 mL of media in 50mL Falcon tubes.
Replace media with the media containing 100 ng/mL nocodazole from step 2 and incubate for 12 h to induce mitotic block.
Very gently remove media and replace with 3mL fresh media (Mitotic shake off - tap flasks against each other 10 times/ Rotate flasks 90 degrees/ tap two flasks against each other 10 times/ Reach 40 times).
Check under the microscope that cells are floating and collect them in 50mL Falcon tubes.
Add 7mL media to each flask, repeat mitotic shake off and add to 50 mL Falcon tubes from step 4.
Spin the Falcon tubes for 3min at 200xg.
Wash pellet with 25mL Dulbecco's Phosphate-Buffered Saline (DPBS).
Spin The Falcon tubes again for 3min at 200xg.
Resuspend in 10mL and plate in 10cm dishes.
Release from mitotic arrest for 120min.
Fix and stain with α-Tubulin antibody to confirm successful synchronization.