Quantification of cancer cell extravasation in vivo.

Cancer cell 'invasiveness' is one of the main driving forces in cancer metastasis, and assays that quantify this key attribute of cancer cells are crucial in cancer metastasis research. The research goal of many laboratories is to elucidate the signaling pathways and effectors that are responsible for cancer cell invasion, but many of these experiments rely on in vitro methods that do not specifically simulate individual steps of the metastatic cascade. Cancer cell extravasation is arguably the most important example of invasion in the metastatic cascade, whereby a single cancer cell undergoes transendothelial migration, forming invasive processes known as invadopodia to mediate translocation of the tumor cell from the vessel lumen into tissue in vivo. We have developed a rapid, reproducible and economical technique to evaluate cancer cell invasiveness by quantifying in vivo rates of cancer cell extravasation in the chorioallantoic membrane (CAM) of chicken embryos. This technique enables the investigator to perform well-powered loss-of-function studies of cancer cell extravasation within 24 h, and it can be used to identify and validate drugs with potential antimetastatic effects that specifically target cancer cell extravasation. A key advantage of this technique over similar assays is that intravascular cancer cells within the capillary bed of the CAM are clearly distinct from extravasated cells, which makes cancer cell extravasation easy to detect. An intermediate level of experience in injections of the chorioallantoic membrane of avian embryos and cell culture techniques is required to carry out the protocol.