CORE--MOLECULAR ANALYSIS

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The purpose of Core C is to provide molecular analysis services to the Project investigators in this Program Project. Western immunoblot, northern and in situ hybridization, and [quantitative, competitive] polymerase chain reaction (RT-PCR) analyses; [DNA fragmentation; electromotive shift assay; enzyme assays for acetylcholine (ACh)-related enzymes and for ICE; and cytokine bioactivity analysis for cytokine cycle proteins] are now available. In addition, analysis of other proteins and their encoding mRNAs that may be of interest (e.g., other cytokines, growth factors, or structural proteins) will be available upon request. S100beta and GFAP ELISAs are available, and other ELISAs can be constructed for rapid screening of large numbers of samples. As a rapid screen for specific mRNAs in cell cultures, micro-in situ hybridization is also available. [The tissue to be analyzed will from human and rodent brain (Projects 1-4), and from cell cultures (Project 3).] Data from analyses performed in this Core will be provided to the Project Investigators as both raw data and preliminary analyzed data. For example, data from northern and western immunoblot analyses and RT- PCR will be returned to the investigator as raw data (pictures of gels and films of transfers) and as ng/mul of extract, relative to total protein, total RNA, G3PDH, or hybridizable ribosomal RNA. Data from ELISAs will be returned as raw data in the form of optical density units per sample, together with a standard curve generated at the same time, and calculation of ng/mug protein and ng/mul extract. Following in situ hybridization, sections or cell cultures will be returned to the investigator or, if requested, sent direction to [project 4] for image analysis. Data from micro-in situ hybridization of cultures will be returned to the investigator as cpm/well relative to total polyadenylated mRNA. [Verification of and values from services performed in this Core will be archived in the free standing computer database in this core.]