The disciplines of Molecular and Cellular Biology have evolved from essentially descriptive and qualitative data analysis to rigorous quantitative determinations of molecular structure and function. In order to stay at the forefront of modern molecular biology, it is absolutely necessary to acquire the ability to quickly and efficiently quantify data and images. This proposal seeks to acquire "state-of-the-art" phosphorimaging capabilities to support ongoing molecular biologic research projects at The Wistar Institute. Each of the projects described herein is aimed at establishing the molecular bases of diseases using modern molecular/cellular biology methods with ultimate goals of improving diagnosis/prognosis and treatment of diseases, such as cancer, AIDS, and other immune-system dysfuncitons and other viral diseases. The molecular targets being examined in these diseases span the entire route of cellular signal transduction: growth-factors, growth-factor receptors and other cell surface receptors, cell adhesion molecules and cytoplasmic structural proteins, soluble kinases and finally nuclear-localized transcription factors.. A comprehensive analysis of these molecules requires sophisticated methods for accurate quantitation of each of the parameters of their function. These parameters include: 1) gene structure, and tpanscriptional activity (via southern, northern and RNAse protection analyses) 2) Tissue distribution of gene activity and developmental expression (via Northern blot and RT-PCR) 3. Protein expression, post- translational modification and detection of associated proteins (via CAT - assay, quantitative in vitro Kinase assays and in vitro transcription. Acquisition of the Phosphorimager will allow each of these parameters to be rigorously quantitated and contribute to each of the research projects described here in a fundamental way.
