Vital cell labelling for the detection of invasive growth in the chick embryo skin invasion assay

Of the various in vitro invasion assays described, only a few use tissues a s substrates, for example. the chick heart fragment assay and the chick emb ryo skin (CES) invasion assay (Noguchi el al., 1978). We have improved cult ure conditions for the CES invasion assay (Schlage, 1989). A sus pension of neoplastically transformed cells is incubated on an explanted piece of ski n from a 9-10-day-old chick embryo. After 1, 2 and 3 days, the explants are fixed and cross-sectioned. As a measure of invasiveness, the number of inv ading cells and their mitotic activity should be evaluated. We tested the s uitability of the vital fluorescence dye PKH-2 (Horan and Slezak, 1989) for improving discrimination between CES cells and invading cells; human cervi cal carcinoma (HeLa), 3-methylcholanthrene (MCA)transformed mouse embryo fi broblasts (10T1/2-MCA7), and MNNG-transformed rat tracheal epithelial cells (RTE-MNNG). The three cell lines formed distinct infiltrates on days 2 and 3, but on day 1, only 10T1/2-MCA7 and R-TE-MNNG cells formed infiltrates. Although PKH-2 labelling was found suitable for the detection of invasion, cellular resolution and dye stability in cryosections is still unsatisfacto ry. To overcome this for routine work, we suggest improving the histologica l processing and using PKH-26, which is a more stable dye. (C) 1999 Elsevie r Science Ltd. An rights reserved.