A range of protocols have been published for making gastruloids, reflecting the ongoing development of this experimental system (in reverse chronological order, [1-4]).
We would direct any researcher to the 2018 protocol available on the Nature Protocol Exchange in the first instance. (Please note that the concentration of N2 supplement is erroneously listed as 4x, when it should in fact be used at 0.5x if making N2B27 medium).
For a visual demonstration of the required techniques, see the article published on the Journal of Visualised Experiments [3], though please note that the order of the wash steps has since changed.
We are continually looking to optimise and refine our gastruloid protocol and will circulate any changes through the Updates page. Please do get in touch if you have any suggestions from your own experience.
Resources Cited:
[1] van Batenburg, V., van den Brink, S.C., Blotenburg, M., Alemany, A., Moris, N., Baillie-Johnson, P., el Azhar, Y., Sonnen, K.F., Martinez Arias, A., van Oudenaarden, A. Generating gastruloids with somite-like structures from mouse embryonic stem cells, 19 February 2020, PROTOCOL (Version 1) available at Protocol Exchange doi: https://doi.org/10.21203/rs.2.18203/v1
[2] Duboule D., Martinez Arias A., Girgin, M., Turner, D.A., Baillie-Johnson, P., Cossy, A.-C., Beccari, L., Moris, N. and Lutolf, M. Generating Gastruloids from Mouse Embryonic Stem Cells, 12 October 2018, PROTOCOL (Version 1) available at Protocol Exchange doi: https://doi.org/10.1038/protex.2018.094
[3] Baillie-Johnson, P., van den Brink, S. C., Balayo, T., Turner, D. A., Martinez Arias, A. Generation of Aggregates of Mouse Embryonic Stem Cells that Show Symmetry Breaking, Polarization and Emergent Collective Behaviour In Vitro. J. Vis. Exp. (105), e53252, doi:10.3791/53252 (2015).
[4] Baillie-Johnson, P., van den Brink, S.C., Balayo, T., Turner, D.A., Martinez Arias, A. Generation of Aggregates of Mouse ES Cells that Show Symmetry Breaking, Polarisation and Emergent Collective Behaviour in vitro. bioRxiv 005215; doi: https://doi.org/10.1101/005215
Peter Baillie-Benson, May 2020