The fast changing field of flow cytometry has required the flow core to replace old instrumentation and up-grade the newer instruments that we have in order to meet the requirements of our CCR investigators. In early FY10 a green laser was added to both the FACSAria SORP and the LSRII SORP to facilitate the sorting and analysis of red fluorescent proteins. Investigators have been busy developing the transgenic mouse models with red fluorescent protiens expressed and have therefore just begun to use the green laser to detect red fluorescent proteins with their gene of interest attached in transgenic mice. Others have found that it detects low expression proteins such as Kir very well with a Phycoerythrin conjugated antibody. Many more of the investigators have increased the complexity of their experiments and are routinely running experiments with 6-9 antibodies to define distinct populations of tumor infiltrating lymphocytes in different tumor models. Various rare event tumor infiltrating lymphocytes have been sorted using 9 antibodies to define the populations in order to detect RNA expression involved in the inflammatory response to the tumor measured by quantitative RT-PCR. The larger number of fluorescence detectors has greatly increased the quality and complexity of information generated about the role of inflammation in tumor progression with each experiment. There should be at least one investigator working on using 12 colors to define their tumor infiltrating lymphocyte populations of interest to determine the cytokines expressed and regulating the inflammation response in tumors. The flow core staff has trained sixteen investigators in the first three quarters of the year to run their own samples teaching the older technology to the more novice in flow cytometry while the more experienced investigators with the need for complex experiments to answer their research questions have been trained on the newer digital instruments with the capability of more antibody combinations. The instruments are used daily by the trained investigators often into the night and on the weekends. This leaves more time for the flow core staff to sort approximately 450 samples and analyze about 7200 samples in the first three quarters of this year. Without the investigators acquiring the data and analyzing their own samples, the government would have to hire one or more probably two full time experienced individuals to perform the same volume of work. At least 6 papers have been published in the past year using data obtained in the flow core. In the past year, data has been generated (or samples have been sorted) for 102 individual scientists from the laboratories of 37 principal investigators. Although the CCR-Frederick Flow Cytometry Core is part of the CIP, investigators from this program account for approximately 47% of the use of this facility. The future goals include keeping up with cutting-edge technology in the flow lab by expanding the pool of investigators trained to run and analyze their own samples to all of CCR in Frederick thereby expanding the use, productivity and quality of the science generated using the core's instrumentation and expertise. This could be achieved by adding a 405 laser to the LSRII Fortessa, replacing the two BD FACScans with small 2 laser 6-color instruments.
| Gudmundsdottir, Bjorg; Gudmundsson, Kristbjorn O; Klarmann, Kimberly D et al. (2018) POGZ Is Required for Silencing Mouse Embryonic ?-like Hemoglobin and Human Fetal Hemoglobin Expression. Cell Rep 23:3236-3248 |
| Thirthagiri, Eswary; Klarmann, Kimberly D; Shukla, Anil K et al. (2016) BRCA2 minor transcript lacking exons 4-7 supports viability in mice and may account for survival of humans with a pathogenic biallelic mutation. Hum Mol Genet 25:1934-1945 |
