Immune escape mechanisms such as the induction/recruitment of immunosuppressive cells and the increased expression of various immunosuppressive molecules including PD-1/PD-1 ligands are essential processes during cancer development and progression, leading to the development of complex immune-suppressive networks in the tumor microenvironment. Thus, high-dimensional immune profiling using CyTOF® technology for single-cell phenotype and function analysis plays an important role to develop more effective cancer immunotherapeutic strategies and to define biomarkers for stratifying responders and non-responders via the detailed analysis of immune responses in cancer patients.

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Cytokine storm is suspected of producing the overzealous immune response driving the severe cardiopulmonary complications that fuel high morbidity and mortality rates in certain COVID-19 infected individuals. IL-1β is one of the key inflammatory cytokines implicated in the cytokine storm syndrome. Inhibition of IL-1β induced cellular signaling with the IL-1 receptor antagonist anakinra has shown promise in treating cytokine storm and is proposed as a potential therapy for subjects with this severe complication of COVID-19. Yet little is known about the immune cell subtypes most responsive to IL-1β induced inflammatory signals in humans. We have developed a custom CyTOF panel and have identified the immune cell targets of IL-1β, and direct and indirect signaling, induced by IL-1β. Our data showing individual heterogeneity in response to IL-1β stimulation and inhibition by anakinra suggest that immune phenotyping of COVID-19 subjects using a similar approach could be effective in identifying those at risk for cytokine storm and those most likely to benefit from IL-1β inhibition.

De Novo Software has been a leading provider of cytometry analysis tools since 1998 and provides data analysis tools for both flow and imaging cytometry. FCS Express 7, De Novo Software’s flagship product, is now being provided with all new Helios purchases to enable Fluidigm users to easily move from acquisition to high dimensional data analysis to report creation in record time. Join us for David’s talk to learn more about the exciting partnership with Fluidigm and to see FCS Express in action.

This presentation will discuss how mass cytometry can benefit vaccine research programs to overcome hurdles, deliver valuable insights and inform the evaluation of candidate vaccines. Results and lessons from the development of a novel vaccine for Coxiella burnetii and other examples will illustrate key benefits, challenge, and considerations for integrating mass cytometry into vaccine research programs.

In line with the role of T cells as critical mediators of the adaptive immune response, recombinational and junctional diversity of the T cell receptor makes each T cell almost entirely unique. Therefore, it makes sense that T cells also display enormous phenotypic and functional diversity in both blood and tissues. This is particularly evident in the context of both infectious disease and cancer, where a great deal of heterogeneity is observed both within individual patient tumors and between patients. A major goal of the lab is to understand the roles of antigen specificity in shaping the heterogeneity of T cells. Here I will talk about our efforts using mass cytometry and single-cell sequencing to identify and profile various antigen-specific T cell populations to gain insights about the boundaries that their specificities place on their capacity to differentiate and take on various phenotypic and functional profiles. For instance, I will describe the implications and hypotheses that come from our recent work in identifying and profiling T cells that are specific for cancer-unrelated antigens and that we designate as bystander T cells. Overall, I hope to highlight the utility of combining broad cellular immune profiling with the analysis of antigen-specific T cells to gain insight about immunological mechanisms of immunity to infectious disease or cancer in humans.

Multiplex immunophenotyping technologies are indispensable for a deeper understanding of biological systems. Until recently, high-dimensional cellular analyses implied the loss of tissue context because most were performed in single-cell suspensions. The advent of Imaging Mass Cytometry™ (IMC™) has enabled highly multiplexed immunohistochemistry of frozen and formalin-fixed, paraffin-embedded (FFPE) tissues using proven CyTOF technology.

This presentation supports the implementation of IMC in other laboratories. I will detail how my team developed an optimized workflow for maximum antibody performance and discuss some of the methodological challenges it needed to overcome to develop a large antibody panel to preserve signal intensity and specificity of antigen detection.

Finally, I will outline why the panel is an excellent immune monitoring tool that can be readily applied in the context of research and clinical trials.

Brain malignancies can either originate from within the CNS (gliomas) or invade from other locations in the body (metastases). A highly immunosuppressive tumor microenvironment (TME) influences brain tumor outgrowth. Whether the TME is predominantly shaped by the CNS micromilieu or by the malignancy itself is unknown, as is the diversity, origin, and function of CNS tumor-associated macrophages (TAMs). Here, we have mapped the leukocyte landscape of brain tumors using high-dimensional single-cell profiling (Cy- TOF). The heterogeneous composition of tissue-resident and invading immune cells within the TME alone permitted a clear distinction between gliomas and brain metastases (BrM). The glioma TME presented pre- dominantly with tissue-resident, reactive microglia, whereas tissue-invading leukocytes accumulated in BrM. Tissue-invading TAMs showed a distinctive signature trajectory, revealing tumor-driven instruction along with contrasting lymphocyte activation and exhaustion. Defining the specific immunological signature of brain tumors can facilitate the rational design of targeted immunotherapy strategies.