High-dimensional cytometry for interrogating immunity and disease — ASN Events
  1. Schedule
  2. Workshop 7: Infection and Immunity 2
  3. High-dimensional cytometry for interrogating immunity and disease

High-dimensional cytometry for interrogating immunity and disease (#70)

Thomas M Ashhurst 1 2 3 4 5 , Paula Niewold 1 5 , Tamara Suprunenko 4 5 6 7 , So Ri S Jung 4 5 6 7 , Zheng L Ling 1 5 , Nicole G Hansbro 8 , Phil M Hansbro 8 , Markus Hofer 4 5 6 7 , Iain Campbell 4 5 6 7 , Adrian L Smith 2 3 5 , Nicholas J. C. King 1 2 3 4 5 7
  1. Viral Immunopathology Laboratory, Pathology, University of Sydney, Sydney, NSW, Australia
  2. Sydney Cytometry Facility, The University of Sydney and Centenary Institute, Sydney, NSW, Australia
  3. Ramaciotti Facility for Human Systems Biology (RFHSB), The University of Sydney and Centenary Institute, Sydney, NSW, Australia
  4. Marie Bashir Institute for Infectious Diseases and Biosecurity (MBI), The University of Sydney, Sydney, NSW, Australia
  5. Charles Perkins Centre, The University of Sydney, Sydney, NSW, Australia
  6. School of Life and Environmental Sciences, The University of Sydney and Centenary Institut, Sydney, NSW, Australia
  7. Bosch Institute, The University of Sydney, Sydney, NSW, Australia
  8. Hunter Medical Research Institute, The University of Newcastle, Newcastle, NSW, Australia

Modern mass cytometry enables characterisation of the immune system by analysing >40 parameters simultaneously on single cells. However, the development of novel fluorescent dyes, and enhancement of acquisition electronics, has led the way in the development of next-generation fluorescence flow cytometers. Because complex interactions between large numbers of individual cells define the behaviour of biological systems, these high-dimensional cytometry platforms provide new tools for interrogating immunity and disease in unprecedented detail. To this end, we deployed two platforms: a Helios mass cytometer, capable of detecting >40 parameters per cell; and a new ‘LSR-X’ SORP platform equipped with 10 high-powered lasers and a 30-parameter detector array. Using these platforms in parallel, we developed closely related mass (40 parameters) or fluorescence (27 fluorescent with 3 physical parameters) cytometry panels to comprehensively map the murine haematopoietic compartment in the bone marrow, spleen, and central nervous system (CNS). Using these panels, we investigated the immune responses in a range of murine disease models, including West Nile virus encephalitis, Zika virus encephalitis, lymphocytic choriomeningitis virus infection/encephalitis, and pulmonary emphysema; in the presence or absence of various genes or therapies. We also incorporated an imaging mass cytometry system, for high-dimensional analysis of CNS tissue sections. Through this approach, we have revealed a systematic and disease-driven re-organisation of haematopoietic output, which enhances the production of cell types that are required in a specific disease scenario. In addition to improving our understanding of disease processes, the simultaneous application of these parallel panels allowed us to perform specific performance comparisons between mass and fluorescence cytometry. Specifically, we compared assay simplicity, acquisition speed, population resolution, and utility in computational analysis. Through this study, we have used high-dimensional cytometry to reveal nuanced and critical changes in the evolution of immune responses, in a range of complex diseases. 

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