? During Phase I of our proposed research, we will develop and validate procedures for recovering, labeling, and analyzing miRNAs from fixed tissue samples. The procedures will be based on the miRNA microarray and fixed tissue RNA isolation systems that we developed in other SBIR-funded programs. The development of our miRNA isolation and labeling procedures will be accomplished using a model system wherein mouse organs will be split and half is flash-frozen and the other half formalin-fixed using a procedure that is commonly employed in hospitals. The frozen and fixed samples will be processed to recover the miRNAs. The miRNAs from the fixed and frozen tissues will be independently labeled and analyzed using miRNA microarrays. The isolation, labeling, and hybridization procedures will be varied until the fixed samples yield the same miRNA expression profiles as the equivalent frozen samples. The fixed sample procedures will) then be used to analyze formalin fixed, human tissue samples to analyze miRNA profiles from multiple organs. The fixed tissue miRNA profiles will be compared to the profiles generated from frozen samples to verify that the fixed tissue miRNA profiling process can be used for stored, human fixed tissue samples. ? During Phase II of our research project, we will use the miRNA isolation, labeling and microarray analysis procedures developed during Phase I to analyze archived, fixed human cancer tissues to identify miRNAs with expression profiles that are significantly different from equivalent, normal tissues. The most interesting miRNAs or miRNA signatures might provide opportunities for diagnostic/prognostic assay development or even an intervention point for therapeutic agents. ? ?
| Szafranska, Anna E; Davison, Timothy S; Shingara, Jaclyn et al. (2008) Accurate molecular characterization of formalin-fixed, paraffin-embedded tissues by microRNA expression profiling. J Mol Diagn 10:415-23 |
| Peltier, Heidi J; Latham, Gary J (2008) Normalization of microRNA expression levels in quantitative RT-PCR assays: identification of suitable reference RNA targets in normal and cancerous human solid tissues. RNA 14:844-52 |
| Doleshal, Martina; Magotra, Amber A; Choudhury, Bhavna et al. (2008) Evaluation and validation of total RNA extraction methods for microRNA expression analyses in formalin-fixed, paraffin-embedded tissues. J Mol Diagn 10:203-11 |
