Figure 8.

EVA-mediated cell aggregation. Nonrecombinant pcDNA.3-transfected CHO and Eva.myc-transfected CHO were detached and labeled, either with PKH26 or PKH2, respectively. After dissociation, the two cell lines were allowed to aggregate in suspension culture, either alone or mixed in a 1:1 ratio, and then analyzed 90 min after incubation in low Ca buffer. Vector-transfected cells are scattered as a single cell suspension (A), whereas large cell clusters are formed by Eva.myc- expressing cells (B). Extent of aggregation is expressed by aggregation index, calculated as D = (N₀ - N₉₀)/N₀. Large aggregates (>5 cells) represented the 8.5% of the total events in mock-transfected cells and 60% in Eva-myc cells (C). In the coculture, EVA-expressing cells aggregate on their own; random incorporation of parental CHO into aggregates never exceeded 10–20% (D). Coaggregation experiments with unlabeled cells were done as described above. Immunofluorescence staining with anti-myc mAb shows selective labeling of cells forming aggregates (E), when compared with phase-contrast analysis of the same field (F). Bars: (A and B) 50 μm; (D–F) 25 μm.