Microfluidic biochips are revolutionizing high-throughput DNA sequencing, immunoassays, and clinical diagnostics. As high-throughput bioassays are mapped to digital microfluidic platforms, the need for design automation techniques is being increasingly felt. Moreover, as most applications of biochips are safety-critical in nature, defect tolerance is an essential system attribute. Several synthesis tools have recently been proposed for the automated design of biochips from the specifications of laboratory protocols. However, only a few of these tools address the problem of defect tolerance. In addition, most of these methods do not consider the problem of droplet routing in microfluidic arrays. These methods typically rely on postsynthesis droplet routing to implement biochemical protocols. Such an approach is not only time consuming, but also imposes an undue burden on the chip user. Postsynthesis droplet routing does not guarantee that feasible droplet pathways can be found for area-constrained biochip layouts; nonroutable fabricated biochips must be discarded. We present a synthesis tool that integrates defect tolerance and droplet routing in the design flow. Droplet routability, defined as the ease with which droplet pathways can be determined, is estimated and integrated in the synthesis procedure. Presynthesis and postsynthesis defect-tolerance methods are also presented. We use a large-scale protein assay as a case study to evaluate the proposed synthesis method.