Cold-temperature plastic resin embedding of liver for DNA- and RNA-based genotyping

The standard practice of tissue fixation in 10% formalin followed by embedd ing in paraffin wax preserves cellular morphology at the expense of availab ility and quality of DNA and RNA. The negative effect on cellular constitue nts results from a combination of extensive cross-linking and strand scissi on of DNA, RNA, and proteins Induced by formaldehyde as well as RNA loss se condary to ubiquitous RNase activity and negative effects of high temperatu re exposure during paraffin melting, microscopic section collection, and ti ssue adherence to glass slides. An effective strategy to correlate cellular phenotype with molecular genotype involves microdissection of tissue secti ons based on specific histopathological features followed by genotyping of minute representative samples for specific underlying molecular alterations . Currently, this approach is Limited to short-length polymerase chain reac tion amplification (<250 bp) of DNA, due to the negative effects of standar d tissue fixation and processing. To overcome this obstacle and permit both cellular morphology and nucleic acid content to be preserved to the fulles t extent, we instituted a system of cold-temperature plastic resin embeddin g based on the use of the water-miscible methyl methacrylate polymer known as Immunobed (Polysciences, Warminster, PA). The system is simple, easy to adapt to clinical practice, and cost-effective. Immunobed tissue sections d emonstrate a cellular appearance equivalent or even superior to that of sta ndard tissue sections. Moreover, thin sectioning (0.5-1.0 mu m thickness) r enders ultrastructural evaluation feasible on plastic-embedded blocks. Tiss ue microdissection is readily performed, yielding high levels of long DNA a nd RNA for genomic and transcription-based correlative molecular analysis. We recommend the use of Immunobed or similar products for use in molecular anatomical pathology.