This Major Research Instrumentation (MRI) Program grant support acquisition of a laser-based flow cytometer at the College of William and Mary, Virginia Institute of Marine Science (VIMS). Partial technician salary support over a two year period will also be supported. The flow cytometer will support a range of marine ecosystem research that will be advanced by the ability to quickly separate abiogneic and biogenic particles and sort individual microbes and even viruses based on characteristics of size, shape and pigment composition as interrogated via laser diffraction and fluorescence from appropriately stained cells. The instrument to be purchased, a BD Influx, will support novel studies of nutrient cycling by marine phytoplankton and heterotrophs by enabling separation of individual microorganisms that can then be cultured and their rates of nutrient cycling measured with stable isotopic labeling techniques. Measurement of species level phytoplankton nitrogen assimilation rates are now possible using combinations of flow cytometry, molecular probes, 15N labeling and mass spectrometry. Other research avenues that will make use of the flow cytometer include marine microbe specific studies of trace metal and nutrient cycling, studies of hydrothermal vent associated benthic invertebrate feeding rates and efficiency, and molecular studies of harmful algal blooms, marine parasites and wastewater effluent borne human pathogens. The cytometer will support regional research use by professors and students from institutions throughout the Commonwealth of Virginia and further afield including faculty at Princeton and the University of Georgia.
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The primary objectives were to purchase a state-of-the-art flow cytometer with fluorescence-activated cell sorting (FACS) capabilities for the Virginia Institute of Marine Science, to train personnel to use the instrument and to establish a core facility at the institute to allow marine scientist students and researchers at VIMS, as well as those from other institutions to use the instrument to further their research activities. Flow cytometers analyze one particle at a time by using a laser to excite the particle with a specific wavelength of light, and then quantify the resulting light scattered by and fluoresced from that particle. These basic light signals are then used to investigate inherent particle characteristics such as size, shape, or pigment composition. Flow cytometers coupled with FACS can be used to identify and define particle characteristics that will allow specific cells of interest to be sorted out of the sample stream for subsequent analysis. VIMS purchased a BD Influx configurable cell sorter produced by BD Biosciences using these funds in the fall of 2012. Two VIMS technicians attended a weeklong training course in California in order to become proficient at using the instrument. In addition, the lead technician attended the BD Art of Sorting: Beyond the Basics course in Feb. 2014. This course covered a wide variety of topics that will allow us to optimize the performance and utility of the Influx. Those topics included the physics of droplet formation and how it affects stream stability, tips for improving sample preparation and viability, gating strategies for sorting and analysis, calculating predicted yield and efficiency using Poisson statistics, techniques for sorting rare populations, small particle sorting, single cell and index sorting, and hands-on practice sorting a variety of cellular samples. The Influx is already being used for several research and environmental monitoring activities at VIMS including studies on the phytoplankton and bacterial communities in local waterways, as well as those in Arctic and Antarctic waters. This instrument is facilitating VIMS scientists’ development of new diagnostic approaches for detecting and quantifying aquatic pathogens of humans and animals marine, and their innovative research in areas as diverse as harmful algal blooms (HABs), microbial community composition analyses including impacts of nutrients and varying temperatures, trace metal biogeochemistry and radiocarbon, to name a few. We are in the process of establishing a VIMS Core Facility that will serve VIMS scientists and those from other institutions. We have determined fees for internal and external scientists to use the instrument on a per week basis, and a website has been created which includes details on the instrument, fee structure information, pictures, and contact information. Note: This Project Outcomes Report for the General Public is displayed verbatim as submitted by the Principal Investigator (PI) for this award. Any opinions, findings, and conclusions or recommendations expressed in this Report are those of the PI and do not necessarily reflect the views of the National Science Foundation; NSF has not approved or endorsed its content.
