Laser capture microdissection-guided fluorescence in situ hybridization and flow cytometric cell cycle analysis of purified nuclei from paraffin sections

Laser capture microdissection (LCM) has recently been identified as a quick , simple, and effective method by which microdissection of complex tissue s pecimens for molecular analysis can be routinely performed. Assessment of g ene copy number by fluorescence in situ hybridization (FISH) is useful for the analysis of molecular genetic alterations in cancer. Unfortunately, the application of FISH to paraffin sections of tumor specimens is fraught wit h technical difficulty and potential artifacts. Our results demonstrate tha t LCM-microdissected nuclei are suitable for FISH gene copy analysis. Ampli fication of genes in cancer specimens can be detected as easily in LCM-prep ared nuclei as in fresh nuclei from cancer tissue specimens. Furthermore, c ontamination of tumor specimens by normal cells can make interpretation of now cytometric cell cycle analysis difficult. Our results show that LCM-mic rodissected nuclei can also be used for now cytometric cell cycle and ploid y analysis. LCM/FISH offers the advantages of multicolor FISH in a morphologically defi ned cell population, without the technical problems of FISH performed on pa raffin sections. This technique should further simplify the methodology req uired to perform copy number analysis of tumor suppressor or protooncogenes in archived cancer specimens. The use of LCM specimens will also improve t he specificity and simplify the interpretation of now cytometric cell cycle and ploidy analysis of breast cancer specimens.