This core will consolidate histology and imaging aspects of the three projects and provide all main technologies in these areas at significant time and cost savings to the program's laboratories. Services provided will include: (1) Specimen collection and preparation: Mouse embryos and placentas, human term placentas from Magee Hospital, cell culture. (2) Processing: Frozen blocks will be processed and cryoembedded by the core. Processing and embedding in paraffin will be outsourced. (3) Tissue archiving: Blocks will be banked to reduce redundancy and decrease future waiting time for specimens that are of common interest to more than one project. The archive will be managed by a digital database. (4) Sectioning: Frozen and paraffin blocks will be sectioned using Dr. Barak laboratory's cryostat (Cryotome FSE, Thermo-Shendon) and fully motorized microtome (Leica RM2255). Extra slides of specimens for multiple uses or shared interest (e.g., detection of multiple probes or antigens for a single project or analyses pertaining to more than one project) will be archived. (5) Staining and labeling: The core will perform histological staining, in situ hybridization, and immunodetection. (6) Imaging: The core will use three existing, camera-fitted bright-field/fluorescent microscopes in Drs. Barak's and Sadovsky's laboratories: An upright Nikon Eclipse E400 with six fluorescent filters for common fluorophores and GFP species, attached to a Qimaging digital color camera;an inverted Zeiss Axiovert 40 CFL microscope with six filters to all GFP species and common fluorphores, fitted with a Nikon DS-QilMc digital B&W camera;and an upright Leitz DM RBE with a standard fluorophore filter set, using alternately the same B&W Nikon camera. Confocal images will be taken as needed at the confocal microscopy core at Magee-Womens Research Institute, equipped with Nikon Al confocal microscope. (7) Reagent consolidation: General buffers, organic solvents, histological stains, blocking agents, in situ hybridization reagents, vectors and riboprobes, primary and secondary antibodies for immunodetection, and detection reagents for common cellular phenomena like proliferation, apoptosis or lipid droplets. This economy- of-scale will reduce redundant purchases of similar reagents. The core will be maintained and operated by a dedicated technician (TBH), who will be trained in all aspects listed above by Drs. Barak, Sadovsky and Chaillet. This technician will report primarily to Dr. Barak and will work both independently on projects assigned directly by each of the three Pis (moderated by Dr. Barak) and with individuals from the three project laboratories, as assigned. This individual will also coordinate maintenance and repair of core equipment. Having a single dedicated technician for this core will centralize and enhance technical proficiencies and avoid prolonged, redundant training of multiple individuals in the same techniques. This strategy will also save at least 0.5 FTE in each individual project, which would have otherwise been dedicated to similar tasks, resulting in a >0.5 FTE reduction for the entire program.
|Shaffer, Ben; McGraw, Serge; Xiao, Siyu C et al. (2015) The DNMT1 intrinsically disordered domain regulates genomic methylation during development. Genetics 199:533-41|
|Larkin, J C; Sears, S B; Sadovsky, Y (2014) The influence of ligand-activated LXR on primary human trophoblasts. Placenta 35:919-24|
|Larkin, Jacob; Chen, Baosheng; Shi, Xiao-Hua et al. (2014) NDRG1 deficiency attenuates fetal growth and the intrauterine response to hypoxic injury. Endocrinology 155:1099-106|
|Mohammadyani, Dariush; Tyurin, Vladimir A; O'Brien, Matthew et al. (2014) Molecular speciation and dynamics of oxidized triacylglycerols in lipid droplets: Mass spectrometry and coarse-grained simulations. Free Radic Biol Med 76:53-60|
|Makkar, A; Mishima, T; Chang, G et al. (2014) Fatty acid binding protein-4 is expressed in the mouse placental labyrinth, yet is dispensable for placental triglyceride accumulation and fetal growth. Placenta 35:802-7|