Crosslinked gelatin beads: A culture method for the study of cell adhesion and invasion

Summary A method is described for culturing invasive cell lines on crosslinked gelatin beads and preparing them for immunocytochemical and morphological observation. Very invasive cells such as Rous sarcoma virus-transformed chicken embryo fibroblasts, and human melanoma LOX and RPMI7951 and breast carcinoma MDA-MB-231 cells will actively degrade this matrix, extending cellular protrusions, called invadopodia, into the sites of degradation. Normal chicken embryo fibroblasts and other non-invasive cell lines do not disrupt the surface of these beads and do not form invadopodia. Invadopodia extending into the bead can be visualized by electron microscopy. Cellular removal of fluorescent fibronectin that has been covalently coupled to the bead surface can be monitored using fluorescence microscopy of frozen-thin-sections. In double label experiments, immunocytochemistry is used to localize antigens in invadopodia at sites of membrane invasion. The materials for bead preparation are inexpensive, and this method has the advantage that many cell types will attach and spread readily on the beads, while only highly invasive cells will invade into the bead.