Facility Details

The CoBRE Redox Molecular Signaling Core is divided functionally and spatially into two sub-cores: the Analytic Redox Biology Sub-Core and the Molecular Signaling Sub-Core. Analytical Redox Biology Sub-Core. The Analytical Redox Biology Sub-Core offers high-quality, accurate measurements of reactive oxygen species, reactive nitrogen species, reactive sulfide species, protein, peptides, metabolites, and lipids in cell culture and tissue samples. High-performance liquid chromatography (HPLC) systems, coupled with UV-vis and fluorescence detectors, are utilized to specifically quantify cellular and mitochondrial superoxide production, hydrogen sulfide pools (free sulfide, sulfide bound to transitional metals, and sulfane sulfur), and thiols (glutathione, GSH/GSSG, cysteine, cystine, homocysteine, persulfides, and glutathionylation). In addition, this core facility employs a versatile and highly sensitive Sievers NO Analyzer (NOA 280i, GE) capable of measuring nitric oxide, nitrite, nitrate, nitrosoheme, and nitrosothiols in a variety of biological samples. Furthermore, an Agilent 8890 GC and 8335 sulfur chemiluminescence detector can provide high-quality data for the identification and quantification of various biological sulfur compounds. To facilitate redox proteomics and metabolomics, this core facility utilizes both a Thermo Scientific™ Orbitrap Exploris™ 480 and a SCIEX Triple Quad 7500 mass spectrometer. The Exploris 480 is a quadrupole-Orbitrap™ mass spectrometer that provides high-resolution, accurate-mass (HRAM) data useful for low-abundance, high-complexity samples at sub-ppb concentrations and/or sub-ppm mass differences. Ideal applications include protein identification and quantification, shotgun proteomics, single-cell proteomics, and tandem mass tag (TMT) multiplexing. The SCIEX Triple Quad 7500 system provides sensitive quantitative and qualitative measurements for metabolomics and lipidomics by MRM scan, production scan, neutral loss scan, enhanced product ion scans, and MRM3 mode.
Molecular Signaling Sub-Core. The Molecular Signaling Sub-Core offers services for molecular cloning and site-directed mutagenesis, as well as design and production of vectors for CRISPR/Cas9 modification of vascular cells. This core provides services for endothelial, smooth muscle, and cardiac myocyte cell isolation, for generation of vascular cell lines, and for lentivirus production for transient or stable modification of cardiovascular cells. In addition, this core provides access to equipment and expertise for studying the cellular effects of hypoxia/reoxygenation injury (Coy Hypoxic Chamber, CLARIOstar Spectrofluorometer), for automated capillary Western blot analysis (ProteinSimple Jess Simple Western), and for high-resolution fluo-respirometry to study mitochondrial function (Oroboros O2k modular system).