In this video, we describe the isolation of metastatic cancer cell colonies from the chicken chorioallantoic membrane (CAM) using shell-less egg culture. This method helps screen for factors that may be responsible for cancer metastasis.
Protocol
All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board. 1. Isolation of Metastatic Colonies At day 5 post injection, remove embryos from the incubator and inspect the embryo CAMs for metastatic colony distribution. Identify the embryos with uniform colony distribution in which compact (or overly invasive) colonies are present. NOTE: The process of cultur…
Representative Results
Figure 1. Representative results for different steps of the screening protocol. (A) Metastatic colonies formed by heterogenous (library transduced cancer cell line HEp3 cells), 5 days post injection. Red arrow shows compact colony (potential positive hit) that should be excised. (B) Metastatic colonies formed by one of the isolated screen hits (<…
Offenlegungen
The authors have nothing to disclose.
Materials
15 mL conical centrifuge tubes.
Corning
CLS430791-500EA
18 gauge x 1 1/2 BD precision
needle
BD
BD305196
we use 1.2mm x 40mm, it is possible
to use shorter needles if preferred
Benchtop centrifuge.
many sources are available
Any TC compatible centrifuge that
can be used to spin down the cells is
suitable
Collagenase
Sigma
C0130-100MG
cotton swabs
many sources are available
must be sterilized before use
Culture media appropriate for the cell
lines used
many sources are available
We grow HT1080, HEp3 and b16 cell
lines in DMEM, 10% FBS media
Egg incubator
many sources are available
An exact model that is necessary
depends on the scale of the screen.
Available sources are MGF Company
Inc., Savannah, GA, or Lyon Electric
Company Inc., Chula Vista, CA
eppendor tubes , 1.5ml
Sigma
T4816-250EA
Fertilized White Leghorn eggs
any local supplyer
fine forceps
many sources are available
must be sterilized begfore use
Image analysis software
We use Nikon Elements
PBS (1x)
many sources are available
Plastic weighting dishes
Simport
CA11006-614
dimensions are 78x78x25mm; many
other sources are available
Metastatic Cancer Cell Colony Isolation from Chicken CAM: A Procedure to Isolate Cancer Cells from Shell-less Egg Cultures. J. Vis. Exp. (Pending Publication), e20470, doi: (2023).