Work flow for performing and analyzing a whole genome RNAi screen for the spreading of Drosophila S2 cells on Con A–coated surfaces. RNAi treatment was preformed in 96-well format (top left), with one unique dsRNA corresponding to one Drosophila gene per well. After 5 d of treatment, cells were plated on Con A and stained for actin, tubulin, and DNA and subjected to high throughput imaging (top right). After acquisition of the image dataset, images were segmented by a watershed transform to identify individual cells (bottom right). A multitude of parameters were extracted from the individual cells (bottom left). For details see Materials and methods. Bars, 10 µm.