Oct 05, 2022
Human induced pluripotent stem cell culture
- 1UCL Institute of Neurology
Protocol Citation: gurvir.virdi 2022. Human induced pluripotent stem cell culture. protocols.io https://dx.doi.org/10.17504/protocols.io.81wgby7dnvpk/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: October 05, 2022
Last Modified: June 01, 2024
Protocol Integer ID: 70857
Keywords: ASAPCRN
Abstract
Cell culture for the maintenance of hiPSCs.
Coat 6 well plates with Geltrex LDEV-Free Reduced Growth Factor Basement Membrane Matrix (Thermo Fisher Scientific), 1:100 dilution for 1 hour before cell passaging.
When hiPSCs are 80-90% confluent, they are passaged:
Old media is aspirated and replaced with 1ml of cell-culture PBS (Gibco, Thermo Fisher Scientific)
PBS is aspirated and replaced with 0.5 mM EDTA (Thermo Fisher Scientific) made up in PBS.
hiPSC are incubated in EDTA for 00:05:00 at 37 °C .
5m
After 5 minutes, the EDTA is aspirated and cells are dissociated with media (E8 or mTesR 1).
hiPSCs are then placed in new 6 well plates at the desired dilution.
All hiPSCs are fed daily with fresh media. All cells were maintained at 37°C and 5% carbon dioxide.