May 14, 2020
- 1Laboratorio Cellule Staminali Post Natali e Terapie Cellulari, IRCCS Istituto Gaslini, Genova, Italy
External link: https://doi.org/10.1371/journal.pone.0244069
Protocol Citation: Marzia Ognibene 2020. Cell migration assay or Transwell assay. protocols.io https://dx.doi.org/10.17504/protocols.io.bgd9js96
Manuscript citation:
Ognibene M, Pezzolo A (2020) Ezrin interacts with the tumor suppressor CHL1 and promotes neuronal differentiation of human neuroblastoma. PLoS ONE 15(12): e0244069. doi: 10.1371/journal.pone.0244069
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: May 14, 2020
Last Modified: May 14, 2020
Protocol Integer ID: 37025
Cells are washed in cold PBS, detached with 5mM EDTA, centrifuged at 1200 g for 7 min and resuspended in serum-free medium.
Transwell chambers (Corning Costar) are prepared filling with 600 μl of complete medium the lower chamber.
The desired amount of cells is plated in 100 μl of serum-free medium in the upper insert (8.0 μm pore size) of each Transwell chamber and incubated at 37°C.
After 24 hours at 37°C, fill with 600 μl of 5mM EDTA an empty well beside each used Transwell chamber.
Eliminate DMEM from the upper insert and move it inside an EDTA well.
Collect DMEM from the lower chamber and put some EDTA inside, to detach cells possibly fallen from the upper insert.
Detach cells migrated on the underside of the upper insert using a plastic scraper and collecting them in the EDTA well.
Mix in one tube cells collected from the lower chamber and scraped from the insert and centrifuge at 1200 g for 7 min, to count the migrated cells.