Figure 4.

Sphere Formation can be Predicted with 98% Accuracy in 24 Hours

For a Figure360 author presentation of Figure 4, see https://doi.org/10.1016/j.isci.2018.08.015.

(A and B) Single cells were seeded and tracked. At 24 hr PreSp were counted and categorized by the number of cells in each. Each count was correlated with eventual sphere formation on days 7 and 14 (for example, there were a total of 153 single-cell MCF-7 PreSp at 24 hr. Of these, 32 eventually formed spheres on day 14, yielding an SFE of 21%). SFE for each size of PreSp is displayed on the y axis. Error bars are weighted SD. (A) Objects over 50 μm were considered spheres for the MCF-7 cell line. (B) Growths over 35 μm on day 7 and 40 μm on day 14 were considered spheres for the T47D cell line. p Values were calculated by Student's two-tailed t test and are displayed above the bars on the graph (N = 3). * Indicates significantly different from all other PreSp sizes with p <.0001.

(C and D) Single cells were again tracked over 14 days in the same manner as in (A) and (B). At 24 hr PreSp sizes were counted. SFEs from (A) were then used in a predictive capacity on these counts to determine how many spheres would form at 14 days. [For example, in one experiment there were 82 two-cell MCF-7 PreSp at 24 hr. The SFE from (A) would predict 62.76 spheres on day 14. The actual spheres formed on day 14 were then compared with these predictions.] The y axis shows the total spheres from three experiments. The white bar is the total prediction calculated using SFEs from previous experiments. The black bar is the total spheres actually observed. Difference between prediction and observation (cumulative error) at 14 days was 2.15% for MCF-7 cells (C) and 2.03% for T47Ds (D). Average error for MCF-7s was 4.47 (±.5) (C) and 2.07 (± 1.71) for T47Ds (D).