Figure 3

Clones generated from a single CD44^posCD24^pos or CD44^posCD24^neg cell possess molecular and functional heterogeneity similar to that of parental line. (a) Clones were generated by sorting single Ca1a CD44^posCD24^pos or CD44^posCD24^neg cells into individual wells of 96-well plates. Only wells confirmed to contain a single cell were expanded. (b) CD44/CD24 expression of representative clones derived from a single CD44^posCD24^neg or CD44^posCD24^pos cell. From the experiment described in (a), seven clones were generated from sorted CD44^posCD24^pos cells and five clones were generated from CD44^posCD24^neg cells. The CD44/CD24 profiles of representative clones are presented. (c) Total RNA was isolated from isogenic CD44^posCD24^neg and CD44^posCD24^pos cells sorted from the clones described in (a, b) and transcript abundance was evaluated via realtime RT-PCR. Data generated with cells sorted from a clone derived from a single CD44^posCD24^neg cell are presented in red. Data generated with cells sorted from a clone derived from a single CD44^posCD24^pos cell are presented in blue. CD44^posCD24^neg (closed bars) and CD44^posCD24^pos (open bars) differences are presented using the delta delta Ct method. Graphs represent means and associated standard errors of three independent experiments. * indicates P < 0.05. (d) Invasion through Matrigel by isogenic CD44^posCD24^neg and CD44^posCD24^poscells freshly sorted from the clones described in (a, b, c). 48 hr following seeding, invaded cells were counted. Graphs represent averages and associated standard errors of experiments performed in triplicate. Data generated with cells sorted from a clone derived from a single CD44^posCD24^neg cell are presented in red. Data generated with cells sorted from clone derived from a single CD44^posCD24^pos cell are presented in blue. * indicates P < 0.05.