Cellular models for siRNA and small molecule high-throughput screening have been widely used in the last decade to identify targets for drug discovery. Here, we present a twofold readout screening approach based on cell viability and multipolar phenotype. To maximize the discovery of potential targets and, at the same time, reduce the number of false positives in our dataset, we have combined focused and rationally designed custom siRNA libraries with small molecule inhibitor libraries. We describe a cellular model for centrosome amplification as an example of how to design and perform a multiple readout screening strategy.