Abstract
Immune responses involve cell interactions within lymphoid and peripheral tissues including tumors. To gain insight into the relationships between immune function and the local tissue environment, we have developed new multiplex 2D (IBEX) and 3D (Ce3D / Ce3D-IBEX) optical imaging pipelines and associated computational methods (RAPID and SPACE) for quantitative, spatially-resolved image analysis.
IBEX is an open source method for iterative imaging used to probe fixed frozen or FFPE mouse and human tissues to as many as 80 parameters at ~ 300 nm lateral resolution. More than 1000 antibody conjugates have been validated for IBEX and a website provides details on these reagents. Ce3D (clearing enhanced 3D) is a fast clearing technique giving excellent signal intensity from directly conjugated antibodies, permitting a high level of multiplexing and IBEX cycling.
Using these methods we have discovered novel commensal-dependent immune clusters at the junction of vagal sensory neurons in normal lungs. In cancers, these tools have revealed how immunotherapy converts suppressive Tregs into type 1 interferon-gamma producing effectors and the unanticipated negative effects of checkpoint therapy on stem-like memory T cells. A method for identifying T cells actively signaling through the antigen receptor has enabled us to identify antigen reactive cells in fixed tissue and to identify the cognate antigen presenting cell. Ce3D has been used to quantify antigen-specific cells in entire lymph nodes, revealing that suppression of autoreactive T cells by Tregs operates by promoting abortive proliferation.
These findings illustrate the power of in situ imaging for probing the molecular, cellular, and spatial aspects of immune responses in situ.
This work was supported by the Intramural Research Program of the NIAID, NIH.
Biosketch
Ronald N. Germain received his M.D. and Ph.D. from Harvard University in 1976. Since then he has investigated basic immunobiology, first on the faculty at Harvard, then at NIAID, NIH. He has contributed to understanding MHC class II molecules, antigen processing, and T cell recognition, more recently pioneering analysis of the immune system using dynamic and static in situ microscopy. He has published more than 400 scholarly research papers and reviews and trained more than 80 fellows. Among other honors, he has been elected to the National Academy of Medicine, the National Academy of Sciences, EMBO, AAAS, the American Academy of Arts and Sciences, named a Distinguished Fellow by the American Association of Immunologists., and designated a Distinguished Investigator by NIH.