The WVU Flow Cytometry & Single Cell Core Facility (FCSCCF) provides instrumentation and scientific support for single cell analysis and sorting. The facility routinely performs analysis of both eukaryotic and prokaryotic cells for expression of intracellular and extracellular proteins, cell cycle, cell proliferation, and cytokine production.

Single and bulk cell sorting is performed based on expression of cell surface antigen(s) and/or expression of genetically engineered intercellular fluorescent proteins. The facility also provides instrumentation and expertise in single cell gene expression profiling,

This facility is currently equipped with four flow cytometers. The Becton-Dickenson (BD) FACSAria III is a 4 laser high-speed bench top sorter equipped with an automated cell deposition unit (ACDU) and housed in a biosafety cabinet for BSL2(+) sorting. The BD LSRFortessa is a 4 laser analyzer equipped with a high throughput sampling module, and a detector to analyze submicron-sized particles. The Cytek Aurora is a full spectrum 3 laser analyzer with a 96 well plate loader. The Luminex Guava easyCyte HT System is a 1 laser analyzer with 96 well plate loader.

For submicron particle analysis, the facility has a Malvern NanoSight NS300 and a Zetasizer Nano Z. The NanoSight NS300 detects, counts, and measure the size of particles. It is equipped with 4 lasers for detection of fluorescently labeled particles. The Zetasizer Nano Z measures the zeta potential (the charge on particles in a solution) via electrophoretic mobility of particles in a solution using laser Doppler microelectrophoresis.

For gene expression analysis, the facility has two instruments, the 10x Genomics Chromium Controller and the Fluidigm C1 Single Cell Auto Prep System. The 10x Genomics Chromium Controller instrument barcodes individual cell’s RNA for bulk single cell RNAseq and multiomic analysis including gene expression, cell surface proteins, antigen specificity, and chromatin accessibility. The Fluidigm C1 Single Cell Auto Prep System uses an integrated fluidic circuit (IFC) chip to isolate 96 individual cells. While in the IFC, RNA/DNA/microRNA is extracted from the single cells followed by amplification and cDNA generation (if needed). The resulting products can be sequenced or used in PCR.

Other instrumentation in the facility include a Miltenyi Biotec gentleMACS™ Otto Dissociator for disassociation of tissues into single cells or homogenization for molecular biology applications, a Miltenyi Biotec AutoMACS™ Pro located in a biosafety cabinet for sterile paramagnetic cell sorting and a Meso Scale Discovery Sector 2400 automated plate reader that detects multiple analytes by electrochemiluminescence.

The facility has auxiliary laboratory space with appropriate equipment to train personnel and support the needs of researchers. The FCSCCF has several software packages including FCS Express, and BD FACSDiva software for flow cytometry data acquisition and analysis. It also has a separate analysis work stations with 2 print quality color laser printers.

The facility is located on the 2nd floor of the Robert C. Byrd Health Sciences Center North building, Rooms 2160 and 2184.

The Transgenic Animal Core Facility (TACF) at WVU offers pronuclear DNA injection for transgenic mouse production, CRISPR/Cas9-mediated genome editing, and embryonic stem cell injection for chimera production. Cryopreservation and rederivation/resuscitation services are also available for the management of genetically modified mice. The core is housed within an SPF barrier facility on the Health Sciences Center campus. Free consultation is also available to assist in construct and CRISPR design, genotyping, colony management, and special husbandry needs.

The Animal Models and Imaging Facility (AMIF) is a full-service shared resource that provides state-of-the-art small animal imaging to West Virginia University (WVU) researchers and their collaborators, as well as extensive training and assistance with planning experiments and interpreting data. . The AMIF currently performs acute and longitudinal bioluminescence and fluorescence studies on mice using a Caliper IVIS Lumina II optical imaging system. In addition, the facility has recently acquired a VisualSonics Vevo 2100 for real time, micro-ultrasound imaging. The AMIF also offers services such as mouse colony management, tissue collection, cryopreservation of sperm, various surgeries and assistance with designing and writing imaging procedures for animal protocols. The AMIF is growing rapidly to support the preclinical imaging needs of researchers at WVU.

The Microscope Imaging Facility provides light microscopy image acquisition and analysis services on a fee-per-use basis. The facility currently has two laser scanning confocal systems, an inverted Zeiss LSM 510 Confocal system with 3 lasers (488, 514, 543 and 633nm) and an upright Zeiss LSM 510 Confocal with multiple lasers/laser lines (405, 458, 477, 488, 514, 543 and 633nm). These systems can do experiments such as 3D rendering and FRAP. The facility has an upright Olympus AX70 epifluorescent microscope equipped with the MicroBrightField Neurolucida and Stereo Investigator software packages. This system is used for color histology records, 3D neuron reconstruction, serial section reconstruction, anatomical mapping and morphometry analysis. For microdissection, the facility has a Zeiss PALM MicroBeam system for non-contact microdissection of single cells or groups of cells. The isolated cells can then be used for DNA or RNA analysis, proteomic analysis or for further cultivation. For live-cell imaging, the facility has a Nikon TE2000S epifluorescent microscope with Prior filter wheels and a Photometrics Coolsnap HQ CCD camera. This microscope also has an Eppendorf FemtoJet microinjection system. This system is run by the MetaMorph and MetaFluor software packages. This system can do multi-color time-lapse, FRET and calcium ratio imaging. For high-end live-cell imaging, the facility has a new Nikon Swept-Field Confocal system with 3 solid-state lasers (491, 561 and 638nm). This system uses a Photometrics QuantEM CCD camera with on-chip gain to maximize signal and to increase the rate of acquisition. This system is also equipped with a laser TIRF illuminator and a high resolution Photometrics Cool-Snap CCD camera. Tissue culture incubators, Bioptechs Delta T4 dish heaters, objective heaters and a micro-perfusion system are available to maintain live cell cultures. An off-line image analysis station is available to the facility users.