Microarray technology has become a standard tool in many laboratories. Agilent Technologies manufactures a variety of catalog and custom long-oligonucleotide (60-mer) microarrays that can be used in multiple two-color microarray applications. Optimized methods and techniques have been developed for two such applications: gene expression profiling and comparative genomic hybridization. Methods for a third technique, location analysis, are evolving rapidly. This chapter outlines current best methods for using Agilent microarrays, provides detailed instructions for the most recently developed techniques, and discusses solutions to common problems encountered with two-color microarrays.