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Principal Investigator: Ferenc Livak
Organization: DIVISION OF BASIC SCIENCES - NCI
Fiscal Year: 2020
Award: $1,192,401
Funding agency: National Cancer Institute
The core is equipped with six analytical instruments: a state-of-the-art BD FACSymphony A5 flow cytometer equipped with five lasers and the capacity to identify 27 fluorochromes also equipped with a high throughput sampler (HTS) attachment that accommodates 96-well plates to further increase the capacity of the instrument. Two high end BD LSRFortessa Special Order Research Product (SORP1 and 2) cytometers with identical configuration, equipped with five lasers and the capacity to identify 18 fluorochromes. One of these instruments, LSRFortessa SORP1 is also equipped with HTS attachment. The core also has one three-laser, digital BD FACSCanto II cytometer and one, 32-channel Sony SA3800 spectral analyzer. In addition, the Core operates a dual-camera, 4-laser ImageStream MarkII imaging flow cytometer equipped with HTS attachment. In FY20 the Core continued to support both basic analytical experiments, in particular by expanding the use of the Sony SA3800 spectral flow cytometer and also advanced, high-dimensional, multi-parameter flow cytometry analysis on the BD FACSymphony A5 analyzer. Several laboratories (Bhandoola, Bosselut, Goldszmid, Trinchieri and Zhuang) begun to use the instrument to expand analysis up to 20-24 color studies. The Core also continued to expand the use of the ImageStream MarkII imaging flow cytometer (Luminex Corp). This instrument records high quality images of every cell in flow and combines the high throughput of flow cytometry with the high information content of microscopic details. It has been used by the Ashwell, Hunter, Taylor, Samelson, Staudt and Yang laboratories among others. The core completed 555 sort requests on four cell sorters in FY20, despite three month of inactivity during the emergency shutdown of NIH. Two BD FACSAria II instruments are equipped with 3-4 lasers with the capacity to simultaneously identify 11-12 fluorochromes. Both of these sorters are enclosed in flexible, BioBubble Benchtop Biocontainment units. One Beckman Coulter MoFlo Astrios EQ instrument, housed in a custom-designed Class II biosafety cabinet, is equipped with five lasers and capable of detecting 19 fluorochromes. One BD FACSAria Fusion cell sorter is also housed in a custom-designed Class II type A2 biosafety cabinet and is equipped with five lasers and has the capacity to identify 18 fluorochromes. These two instruments have configurations similar to the two high-end LSRFortessa SORP flow cytometers. Further, the two Class II biosafety cabinets and two BioBubble Benchtop Biocontainment units, combined with the separate housing in a dedicated laboratory space in Room 6008A allow the LGI FC37 to operate in full compliance with NIH Policy for Biosafety of Cell Sorters (July 28, 2012), to perform all sorts at a BSL-2 with enhanced precautions aerosol containment level. In FY20 the Core in a joint effort with the NCI Single Cell Analysis Facility (SCAF) in BG37, supported the testing and purchase of a new, Sony MA900 cell sorter. This instrument is housed in the SCAF laboratory but operated by LGI FC37 personnel and is specifically used for 96-/384-well single cell sorting and for pre-enrichment of samples for 10xChromium single cell sequencing applications. This collaboration between the LGI FC37 and SCAF is further evidence to the strategic vision of the two services in recognizing the rapid advance in single cell analysis platforms. The LGI FC37 served 368 users from 82 CCR laboratories as well as from 4 additional laboratories of three other NIH institutes in FY20. Several of these laboratories heavily depend on the services of the Core in conducting high-dimensional analyses aimed at better understanding of lymphocyte development (Ashwell, Bosselut (LICB] and Bhandoola [LGI] labs) and tumor-associated immune responses (Trinchieri lab [CIP] and Zhuang lab [NOB]). Single cell sorting helps the rapid generation of CRISPR-induced mutant cell lines (Hunter [LCBG], Lal [CGB], Nussenzweig [LGI] labs among others) and is becoming an essential tool in single cell genomic analyses (Aldape lab [LP], Bosselut lab [LICB], Merlino and Yang labs [LCBG]). The LGI FC37 provides critical help to the initial phases of translational research into lymphoma targeting screens (Staudt lab [LMBr]), glioblastoma research (Gilbert, Chunzhang Yang [NOB] labs) and tumor-specific immune responses (Waldman lab [LMBR] and Hodge lab [LTIB]). The LGI FC37 has trained 102 new users in FY20 on analytical instruments and has also trained one new user for cell sorting. New users are instructed to review instrument and software training tutorials before attending a 4-hor hands-on training session that includes startup and shut down procedures for the cytometers, setting up experiments, basic instrument troubleshooting, and data analysis. The Core staff provides follow-up assistance and helps new users develop confidence in using the technology correctly. Cell sorter training is offered to select users who need frequent, possibly after hour sorting time. The LGI FC37 employs an entirely online scheduling and record keeping platform using the NIH LabShare application. Appropriate policies are in place to ensure safe, fair and equitable access of Core resources to all registered users. On March 23, 2020 in compliance with NCI/NIH emergency orders, the FC37 entered minimum maintenance operations due to the COVID-19 pandemic. The Core continued to provide limited assistance and instrumentation for analytical work of CCR-exempted, COVID-19-related research. Since June 22, the first phase of reopening of NIH, the Core has been operating at approximately 50% capacity, with two staff members on site for three days a week and offering a 6-day/week support. Analyzer and cell sorter support has been restored to the extent that is consistent with the NIH social distancing guidelines currently in effect on the Bethesda campus. During the minimal maintenance period, Core staff communicated via online meeting tools, participated in 6 flow cytometry journal clubs and organized a technology meeting with Kinetic River, Inc. The LGI FC37 also initiated several meetings with other NCI flow cytometry cores and services and lead the development of joint policy proposals on handling potentially SarsCoV-2-infected samples and principles of safe operations of shared services upon reopening. Despite these challenges, the LGI FC37 remains committed to be a leader of flow cytometry services at CCR by offering the highest quality, reliable support to the largest number of NCI investigators on campus The LGI FC37 is dedicated to the introduction of innovative flow cytometry-based technologies to further advance the cutting-edge cancer research conducted at NIH.
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