By D. Rusciano, D.R. Welch and M.M. Burger (Eds.)
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Additional info for Cancer Metastasis: In vitro and in vivo Experimental Approaches
This gentle washing process is repeated four times. Once the matrix is free of cells, plates are rinsed with PBS at least three times. Lyric removal of EC is usually performed in two steps. 05% BSA can be added) 15 min at 37~ and then with detergent (either Ch. 2 HOMOTYPIC AND HETEROTYPICCELLADHESIONIN METASTASIS 55 Fig. 2. Positive and negative adhesion of fibroblasts on fibronectin and tenascin coating of a tissue culture dish, after saturation with BSA. After 1 h of plating in serum-free medium, cells appear loosely attached on BSA, well spread on fibronectin and not attached on tenascin.
1995). Along this line, Pauli and Lee (1988) have shown that components of the ECM can modulate EC derived from large vessels (such as aorta or umbilical vein) to assume phenotypic traits of the specific microvascular endothelium of the organ from which the organ-specific biomatrix was prepared. Using monolayers of thus modulated bovine aortic EC (BAEC) in a classical adhesion assay, they show that tumor cells which metastasize to a given organ have 46 CANCER METASTASIS a significantly higher binding affinity for BAEC grown on ECMs of the preferred, metastasized organ, than they have for BAEC grown on ECMs of other organs not targeted by these tumor cells.
A further centrifugation in a discontinuous sucrose density gradient (5, 20 and 30% sucrose in PBS) yields single organ cells in the 20% layer; erytrocytes, platelets and membrane debris in the 5% layer; and some aggregates in the 30% layer. Sucrose is finally removed from the single organ cell suspension by centrifugation in PBS, leaving less than 5% erythrocytes and 7% phagocytic cells contaminating the suspension. If further removal of aggregates is required, the suspension can be passed through a cell strainer.
Cancer Metastasis: In vitro and in vivo Experimental Approaches by D. Rusciano, D.R. Welch and M.M. Burger (Eds.)