3D Organoids from primary melanoma cell lines and from iPSc-derived neural crest stem cells

Scope of the method

The Method relates to
  • Human health
The Method is situated in
  • Basic Research
Type of method
  • In vitro - Ex vivo
This method makes use of
  • Human derived cells / tissues / organs
Specify the type of cells/tissues/organs
Human iPSC; Primary melanoma cells


Method keywords
  • 3D culture
  • organoid
  • single-cell
  • ECM
  • hydrogel
  • PEG
Scientific area keywords
  • melanoma
  • enhancer
  • single-cell RNA seq
  • single-cell ATAC seq
Method description

We propose to generate three-dimensional tumoroids from the primary melanoma cell lines, as well as 3D organoids from the iPSc-derived neural crest stem cells. We will use the AggreWell system (STEMCELL Technologies) to generate uniform, size-controlled three-dimensional spheroids. After 5 days in the AggreWell plate, the spheroids are moved to a PEG-based artificial ECM hydrogel (Gjorevski et al.; Nature Protocols 2017). The organoids can be cultured for weeks in these PEG-droplets. At different time points during organoid culture, organoids will be used for immunostaining and/or for single-cell sequencing. We will dissociate the PEG gel to obtain single cells by use of the cell-dissociation enzyme TrypLE.

Lab equipment
  • Biosafety cabinet ;
  • Cell incubator CO2-connected ;
  • Centrifuge for plates.
Method status
  • Published in peer reviewed journal

Pros, cons & Future potential


3D organoids mimic tissue architecture heterogenous cell culture to study cellular differentiation enhancer testing.


Not every cell type/tissue can be studied.


Different cell types are studied to form organoids.

Future & Other applications

Drug application: concentration and activity can be tested.

References, associated documents and other information


Gjorevski N & Lutolf MP. Synthesis and characterization of well-defined hydrogel matrices and their application to intestinal stem cell and organoid culture. Nature Protocols 12 (11); 2263-2274 (2017)

Contact person

Valerie Christiaens


VIB - KU Leuven
Department of Brain and Disease Rearch
Lab of Computational Biology
Flemish Region