Sep 23, 2023

Cell culture

DOI
https://dx.doi.org/10.17504/protocols.io.n2bvj3y5blk5/v1
  • 1Sascha Martens lab, University of Vienna, Max Perutz Labs - Vienna
Elias Adriaenssens
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DOI: https://dx.doi.org/10.17504/protocols.io.n2bvj3y5blk5/v1
Protocol Citation: Elias Adriaenssens 2023. Cell culture. protocols.io https://dx.doi.org/10.17504/protocols.io.n2bvj3y5blk5/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: July 07, 2023
Last Modified: May 31, 2024
Protocol  Integer ID: 84663
Keywords: Cell culture, ASAPCRN, routine cell culture maintenance, cell culture this protocol, cell culture, cell, maintenance
Funders Acknowledgements:
Aligning Science Across Parkinson’s (ASAP)
Grant ID: ASAP-000350
Marie Skłodowska-Curie MSCA Postdoctoral fellowship
Grant ID: 101062916
Abstract
This protocol describes routine cell culture maintenance.
Materials
Reagents
  • Dulbecco Modified Eagle Medium (DMEM, Thermo Fisher)
  • Fetal Bovine Serum (FBS, Thermo Fisher)
  • HEPES (Thermo Fisher)
  • non-essential amino acids (NEAA, Thermo Fisher)
  • Penicillin-Streptomycin (Thermo Fisher)
  • Iscove’s Modified Dulbecco’s Medium (IMDM, Thermo Fisher)

Cell culture
5m
Grow HeLa and HEK293T cells in Dulbecco Modified Eagle Medium (DMEM, Thermo Fisher) supplemented with 10% (v/v) Fetal Bovine Serum (FBS, Thermo Fisher), 25 millimolar (mM) HEPES (Thermo Fisher), 1% (v/v) non-essenMal amino acids (NEAA, Thermo Fisher), and 1% (v/v) Penicillin-Streptomycin (Thermo Fisher).
Culture HAP1 cells in Iscove’s Modified Dulbecco’s Medium (IMDM, Thermo Fisher), supplemented with 10% (v/v) Fetal Bovine Serum (FBS, Thermo Fisher) and 1% (v/v) Penicillin-Streptomycin (Thermo Fisher).
Regularly test all cell lines for mycoplasma contaminations.
Split the cells by trypsinization once confluent.
To this end, remove medium with Pasteur pipette.
Wash cells with 1x PBS and then add trypsin for 00:05:00 at 37 °C .
5m
Afterwards, resuspend cells in medium and retain 10% for further culturing and expansion.
Discard the rest or collect for seeding for experiments.