The advent of high-density complementary DNA microarray technology, with its capacity for simultaneous monitoring of thousands of genes, provides a unique opportunity for high-throughput expression analysis of cancer. Although most current microarray studies have been performed with genetic material obtained in vitro, a major advance in functional genomic investigations would be the ability to perform array-based expression analysis with genetic material obtained in vivo and originating from morphologically distinct cellular populations, including various stages of cancer progression. Until recently, in vivo analysis of tumor-specific genomic alterations, array-based or otherwise, has been hampered by the inability to obtain specific cell types accurately from cancerous tissue. The recent development of laser capture microdissection allows for accurate and rapid procurement of specific cell populations within complex tissue and provides the opportunity to perform array-based expression profiling of genetic material obtained in vivo. We describe the combined use of laser capture microdissection and high-throughput complementary DNA microarrays to monitor gene expression levels in 13 breast cancer specimens. By using different data visualization tools (clustering and multidimensional scaling) we demonstrate that expression profiles of more than 8,000 genes can be successfully generated using non-amplified RNA derived from distinct cell populations within several different morphological stages of human breast cancer.