Immunoassay and Imaging Core

IMMUNOASSAY AND IMAGING CORE: PROJECT SUMMARY Key correlates of immune protection against viral pathogens include virus specific memory T cells resident in tissues and high affinity virus-neutralizing antibodies circulating in blood. During a primary immune response, newly encountered antigen is transported from the sites of exposure to regional lymphoid tissue where naive lymphocytes encounter antigen-presenting dendritic cells and macrophages which critically regulate the priming, expansion and activation of antigen specific T and B cells. Memory T cells derive from activated or effector T cells generated during the primary immune response and differentiate into circulating, tissue surveilling memory T cells as well as tissue resident memory T cells (TRM) that are retained in diverse tissue sites. Immune protection at barrier tissue sites – the primary point of pathogen invasion such as lung - is mediated by TRM established by previous antigen exposures that are specially adapted to persist in the tissue microenvironment and provide a highly localized defense against pathogen re-entry. Long lasting humoral immunity is established via selection of somatically hypermutated B cell antigen receptors in the germinal centers of lymphoid tissue – a process dependent on help from CD4 T follicular helper cells. Thus development and maintenance of cellular and humoral immunity is critically dependent on immune cell interactions in lymphoid and non-lymphoid barrier tissues. This service core will provide quantitative assays of the cellular and humoral arms of the immune response using in situ multiplex staining to define key T cell and dendritic cell populations mediating immune protection in tissues and serological assays to define humoral anti-SARS-CoV- 2 and anti-influenza antibody responses to infection and vaccination. The Aims of the service core are 1) Perform large scale quantitative spatial analysis of immune cells within human tissues; and 2) Provide quantitative and functional analysis of anti-viral antibodies in human serum. For Aim 1, molecular signatures obtained by single cell sequencing of antigen-specific T cells (Project 1) and dendritic cells (Project 2) in human tissues will be used to build multiplex imaging panels to visualize these cells in-situ. The core will support tissue preservation, multiplex staining and analysis of multiplex imaging with a focus on identifying novel antigen-specific TRM and dendritic cell populations across multiple lymphoid and barrier tissue sites and donors. These studies will complement detailed molecular signatures of tissue immune cells with information about their spatial context and microenvironment. For Aim 2, assays of circulating virus specific antibodies will be used to assess SARS-CoV-2 immune protection and vaccination status in organ donors for correlation with tissue immune responses and other donor-specific covariates (Project 1) and to assess SARS-CoV-2 and influenza vaccine responses longitudinally in multiple sclerosis patients taking immunosuppressive medications (Project 3). Together, data generated from this core, in support of the overall program goals, will help define factors that shape tissue and humoral immune protection in humans.