Flow sorting in the study of cell-cell interaction

Undifferentiated mouse teratocarcinoma cells were cocultivated with differentiated mouse endoderm cells in order to study the possible induction of teratocarcinoma cell differentiation. A difference in DNA content between the two cell types was experimentally introduced to enable the reisolation of the teratocarcinoma cells after cocultivation. Pseudotetraploid (2s) endoderm cell lines were produced from pseudodiploid (1s) cells by treatment of these cells with cytochalasin B and flow sorting of tetraploid cells, using Hoechst 33342 as a viable DNA stain, with subsequent cloning of sorted single cells. In model experiments, where mixtures of 1s teratocarcinoma and 2s endoderm cells were stained with Hoechst 33342, the teratocarcinoma cells could be reisolated with a purity of about 97%. After a cocultivation period of 24 days viable teratocarcinoma cells could be isolated from the cocultivation mixture with a purity of 95%. Two dimensional analysis of the protein pattern of these cells indicated that cocultivation did not induce a differentiated (endoderm) pattern. Therefore according to this analysis the teratocarcinoma cells were not induced to differentiate during a 24 day cocultivation period. The method described offers excellent possibilities for studying cell-cell interaction in vitro.