Tissue functionality is determined by the characteristics of tissue-resident cells and their interactions within their microenvironment. Imaging Mass Cytometry offers the opportunity to distinguish cell types with high precision and link them to their spatial location in intact tissues at sub-cellular resolution. This technology produces large amounts of spatially-resolved high-dimensional data, which constitutes a serious challenge for the data analysis. We present an interactive visual analysis workflow for the end-to-end analysis of Imaging Mass Cytometry data that was developed in close collaboration with domain expert partners. We implemented the presented workflow in an interactive visual analysis tool; ImaCytE. Our workflow is designed to allow the user to discriminate cell types according to their protein expression profiles and analyze their cellular microenvironments, aiding in the formulation or verification of hypotheses on tissue architecture and function. Finally, we show the effectiveness of our workflow and ImaCytE through a case study performed by a collaborating specialist.

MOTIVATION: High-dimensional mass cytometry (CyTOF) allows the simultaneous measurement of multiple cellular markers at single-cell level, providing a comprehensive view of cell compositions. However, the power of CyTOF to explore the full heterogeneity of a biological sample at the single-cell level is currently limited by the number of markers measured simultaneously on a single panel. RESULTS: To extend the number of markers per cell, we propose an in silico method to integrate CyTOF datasets measured using multiple panels that share a set of markers. Additionally, we present an approach to select the most informative markers from an existing CyTOF dataset to be used as a shared marker set between panels. We demonstrate the feasibility of our methods by evaluating the quality of clustering and neighborhood preservation of the integrated dataset, on two public CyTOF datasets. We illustrate that by computationally extending the number of markers we can further untangle the heterogeneity of mass cytometry data, including rare cell-population detection. AVAILABILITY AND IMPLEMENTATION: Implementation is available on GitHub (https://github.com/tabdelaal/CyTOFmerge). SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Single-cell analysis through mass cytometry has become an increasingly important tool for immunologists to study the immune system in health and disease. Mass cytometry creates a high-dimensional description vector for single cells by time-of-flight measurement. Recently, t-Distributed Stochastic Neighborhood Embedding (t-SNE) has emerged as one of the state-of-the-art techniques for the visualization and exploration of single-cell data. Ever increasing amounts of data lead to the adoption of Hierarchical Stochastic Neighborhood Embedding (HSNE), enabling the hierarchical representation of the data. Here, the hierarchy is explored selectively by the analyst, who can request more and more detail in areas of interest. Such hierarchies are usually explored by visualizing disconnected plots of selections in different levels of the hierarchy. This poses problems for navigation, by imposing a high cognitive load on the analyst. In this work, we present an interactive summary-visualization to tackle this problem. CyteGuide guides the analyst through the exploration of hierarchically represented single-cell data, and provides a complete overview of the current state of the analysis. We conducted a two-phase user study with domain experts that use HSNE for data exploration. We first studied their problems with their current workflow using HSNE and the requirements to ease this workflow in a field study. These requirements have been the basis for our visual design. In the second phase, we verified our proposed solution in a user evaluation.

Abstract To understand how the immune system works, one needs to have a clear picture of its cellular compositon and the cells' corresponding properties and functionality. Mass cytometry is a novel technique to determine the properties of single‐cells with unprecedented detail. This amount of detail allows for much finer differentiation but also comes at the cost of more complex analysis. In this work, we present Cytosplore, implementing an interactive workflow to analyze mass cytometry data in an integrated system, providing multiple linked views, showing different levels of detail and enabling the rapid definition of known and unknown cell types. Cytosplore handles millions of cells, each represented as a high‐dimensional data point, facilitates hypothesis generation and confirmation, and provides a significant speed up of the current workflow. We show the effectiveness of Cytosplore in a case study evaluation.