Background

We initially designed this panel for the analysis of the PBMCs from patients who have undergone alloHSCT, particularly those enrolled in drug studies for the prevention and treatment of GVHD. Prior studies in humans and animal models have implicated many immune cell types in the initiation and progression of GVHD, and the data have, at times, conflicted, depending on species, model, and individual laboratories. In particular, prior studies have identified imbalances in T regulatory cells (Tregs), T follicular helper (Tfh) cells, T helper type 1 (Th1), type 2 (Th2), type 17 (Th17), myeloid derived suppressor cells (MDSCs), natural killer cells (NKs), dendritic cells (DCs), and others in modulating engraftment, GVHD, and treatment responses [5, 6]. Accordingly, we aimed to develop a standardized panel to capture all major human lymphoid and myeloid populations with deep T cell phenotyping in a single analysis, thus reducing experimental variability, redundancy, and the need for a high quantity of input cells. As to the last point, post-HSCT patients typically have few circulating leukocytes until hematopoietic engraftment and reconstitution. Thus, multiple flow cytometry panels and/or CyTOF analyses pose a greater challenge than a single, comprehensive flow-based panel. Beyond our HSCT-focused studies, this panel should find broad application in the study of many inflammatory and neoplastic conditions. Of note, this panel uses antibodies targeting exclusively surface receptors, making fixation and permeabilization unnecessary.