Grade II astrocytoma is defined as a low-grade tumor, yet patients have a wide range of survival and tumors can quickly progress to high-grade astrocytoma/glioblastoma. Previous studies using comparative genomic hybridization (CGH) failed to demonstrate frequent copy number aberrations (CNA) in these tumors. This may be related to technical difficulties because infiltrating astrocytic tumors are often intermixed with normal brain tissue. We developed methods to exclude most normal tissue and use small amounts of DNA for CGH by microdissecting small regions of tumor from paraffin sections and amplifying extracted DNA using degenerate oligonucleotide-primed polymerase chain reaction (DOP-PCR). Using this method, we examined 30 grade II astrocytoma cases. We found CNA in 25 cases (83%), with a mean of two CNA per case. The most frequent CNA were gains on 7q (12 cases), 5p (5 cases), 9 (5 cases), and 19p (3 cases), and losses on 19q (7 cases), 1p (6 cases), and Xp (3 cases). Gain on 7q and losses on 1p/19q were mutually exclusive. This is the first report on the genetic characterization of low-grade astrocytomas using CGH from microdissected and formalin-fixed tissue. The comparatively large number of cases in this study allows us to suggest that these tumors are genetically subgrouped.