Next-generation DNA sequencing has revolutionized the discovery of rare polymorphisms, structural variants, and novel transcripts. To meet the demand for fast, cost-effective, and accurate genome analysis methods from small scale studies to large sample cohorts, Agilent Technologies has developed the SureSelect™ Target Enrichment System. Available for the Illumina, SOLiD, and 454 NGS sequencing platforms, SureSelect is a highly robust, customizable, and scalable system that focuses analyses on specific genomic loci by in-solution hybrid capture. In addition, Agilent has introduced SureSelect XT for Illumina and SOLiD, which combines gDNA prep, library prep, and SureSelect Target Enrichment reagents in one complete kit. Both SureSelect and SureSelect XT demonstrate high performance, as measured by capture efficiency, uniformity, reproducibility, and SNP detection. We highlight the utility of the SureSelect system across a wide range of target sizes and genome complexity using pre-designed catalog libraries targeting cancer gene sets, sequences encoding the kinome, and both human and mouse All Exon content. In addition, user-defined custom content can be easily developed using the Agilent eArray software with candidate variant coordinates as input. User-defined content can be manufactured on-demand as a custom SureSelect kit, or combined with pre-defined Agilent catalog content using the Plus option. We propose a novel approach for variant discovery - using SureSelect catalog designs to uncover candidate variants, followed by the design of smaller focused custom libraries for SNP validation and region profiling. By pooling many samples together per lane or slide, SureSelect multiplexing kits for Illumina and SOLiD enable validation across large sample cohorts with substantial cost savings. Accurate post target enrichment pooling is facilitated by the Agilent Bioanalyzer and QPCR NGS Library Quantification kits which ensure equal representation across samples. Further efficiencies are realized using the Bravo Automated Liquid Handling Platform to meet the need for parallel preparation of multiplexed libraries.