Intravenous Injection of Cancer Cells into Chicken Chorioallantoic Membrane Vasculature: A Procedure to Establish Chorioallantoic Membrane Cancer Model to Study Cancer Invasion and Metastasis

Published: April 30, 2023

Abstract

Source: Stoletov, K. et al. Discovery of Metastatic Regulators Using a Rapid and Quantitative Intravital Chick Chorioallantoic Membrane Model. J. Vis. Exp. (2021)

This video demonstrates the procedure of intravenously injecting cancer cells into the chicken chorioallantoic membrane vasculature to generate a CAM cancer metastasis model. The injected cells eventually form scattered single-cancer cell colonies, allowing for the study of cancer invasion and metastasis.

Protocol

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board. 1. Preparation of the Cancer Cells for Injection NOTE: Metastatic colony selection is based on a phenotype established by fluorescent cancer cells, derived from an expression library or vector tagged with fluorescent protein such as green or red fluorescent protein (see the whole screen flow …

Representative Results

Figure 1: Outline of cancer cell injection and metastatic colony isolation. (A) Flowchart outlining the chicken embryo screening platform steps. (B) Cancer cell injection set up on the stereo fluorescent microscope stage. (C) Injection of the cancer cells into the CAM vasculature. (D) Image showing successful can…

Divulgaciones

The authors have nothing to disclose.

Materials

1 mL disposable syringes BD BD309659
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
2.5% Trypsin solution many sources are available
4T1 mouse breast cancer ATCC CRL-2539
B16F10 mouse melanoma cell line ATCC CRL-6475
Benchtop centrifuge Many sources are available Any TC compatible centrifuge that can be used to spin down the cells is suitable
Circular coverslips, 22 mm Fisher Scientific 12-545-101
Collagenase Sigma C0130-100MG
Confocal microscope We use Nikon A1r
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
HT1080 human fibrosarcoma cell line ATCC CCL-121
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 supplier
Plastic weighting dishes   Simport  CA11006-614 Dimensions are 78x78x25mm; many other sources are available
Square petri dishes (used as lids for the weighting dishes)   VWR CA25378-115  Dimensions are 100x100x15mm; many other sources are available
Stereo fluorescent microscope  We use Zeiss Lumar v12
Sodium borosilicate glass capillary tubes, outer diameter 1.0 mm, inner diameter 0.58 mm, 10 cm length   Sutter Instrument BF100-58-10
Tygon R-3603 laboratory tubing   Cole-Parmer AAC00001  1/32 in inner diameter, 3/32 in. outer diameter, 1/32 in. wall thickness
Fine forceps Many sources are available Must be sterilized before use
Hemocytometer  Millipore-Sigma MDH-4N1-50PK

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Intravenous Injection of Cancer Cells into Chicken Chorioallantoic Membrane Vasculature: A Procedure to Establish Chorioallantoic Membrane Cancer Model to Study Cancer Invasion and Metastasis. J. Vis. Exp. (Pending Publication), e20469, doi: (2023).

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