Model
Spheroids
Proliferation
Flow cytometry
Fluorescent
imaging
Co-culture assays
Cell migration
Cell invasion
Flow adhesion
Tubule
formation
What is it and what is it used for?
3D ball of cells, often tumour cells, used to study 3D
properties of the cells in response to treatment or in
interaction with other cell types e.g. macrophages.
Often used for implantation into animal models.
Three methods commonly used:
 Cell counting using trypan blue exclusion,
manually or
 MTS assay – indirect measure of proliferation
as it measures activity of a mitochondrial
dehydrogenase enzyme
 BrdU assay – measures incorporation of BrdU
into the DNA, however, this could also occur
during DNA repair
Can be used to study expression and change in
expression of cell surface and inter-cellular proteins.
Can look at multiple proteins at once (multi-flourescent
channels)
Used for sub-cellular localisation of proteins and
assessing changes in cellular architecture e.g.
cytoskeletal changes
Can be done in 2D (e.g. fibroblasts and endothelial cells
to form endothelial tubules) or in 3D (e.g. 2 or more cell
types in spheroids) to investigate interactions between
cell types in response to treatment.
Scratch assay – confluent monolayer of cells with a
‘scratch’ made using a pipette or by having a gate
preventing cells growing in a set area. The cells then
migrate into the region (proliferation inhibited by
Mitomycin C) and the rate of gap closure is assessed.
Boyden chamber – cells migrate through a porous
membrane towards a chemoattractant.
These are both used to assess migration in response to
treatment
Similar to Boyden chamber but uses transwell chambers
which can be coated with a monolayer of cells or a layer
of Matrigel and cells invade across this.
Cells such as endothelial cells grown in a monolayer
then placed in a flow chamber. Tumour cells are then
passed across the top under flow conditions and the
number of tumour cells that adhere to the endothelial
cells are assessed. This is currently being adapted to
investigate invasion under flow as well.
Matrigel assay – endothelial cells placed on Matrigel
form tube like structures over the course of 6-24 hours.
These ‘tubules’ do not contain lumen.
EC spheroid assay – The ECs sprout out from spheroids
suspended in collagen in the presence of VEGF. Difficult
to analyse accurately.
These are used to assess angiogenic properties and
responses in vitro.
Who does it?
Claire Lewis
Gill Tozer
Ingunn Holen
Gill Tozer
Carolyn Staton
Peter Grabowski
Carolyn Staton
Craig Murdoch
Sue Newton
Spencer Collis
Chryso Kanthou
Peter Grabowski
Chryso Kanthou
Claire Lewis
Carolyn Staton
Carolyn Staton
Chryso Kanthou
Carolyn Staton
Karen Sisley
Carolyn Staton
Karen Sisley
Ingunn Holen
Craig Murdoch
Munitta Muthana
Carolyn Staton
Chryso Kanthou