The strategy is to begin by defining the simplest EPs that are well characterized (e.g., CCR7) and work toward the more complex EPs that are less characterized. Similar to the need for biological knowledge necessary for the interpretation of traditional gating analysis, the use of a biological reference point gives context to analysis of the modeled data. In the model, the events are distributed equally across the states for each EP, whether it is considered alone or in concert with other markers. Therefore, the analysis can be

approached one measurement at a time, allowing for a scalable analysis method to a high-dimensional set of measurements, including Oligomycin A supplier unknown elements. Additionally, in traditional gating, overlaps in populations require subjective gating decisions. Flow cytometry standardization studies have identified gating as the largest component in variability of results between laboratories (Jaimes et al., 2011 and Maecker et al., 2005). In PSM, regions defined along a progression axis can automatically account for population overlaps. Many studies have demonstrated the link between phenotypic expression markers on CD8+ T cells with functional properties, including ex vivo effector function. (Appay et al., 2008, Hamann et al., 1997, Lefrancois

and Obar, 2010 and Sallusto et al., 1999). With these observations, much research click here has focused on the classification of effector and memory T-cell subpopulations and their respective functions. The phenotypic heterogeneity in memory T-cell populations has confounded the definition of an accepted Etofibrate model describing immunological development of CD8+ T cells. To approach the classification of memory/effector

subpopulations from a new angle, PSM was applied to healthy donors’ PBMCs stained with CD8+ T-cell markers. The progression plots show three major transitions forming four stages based on CD45RA and CD28, where changes in marker intensities presumably reflect the changes in functional states. This analysis of CD8+ T-cell differentiation is somewhat in contrast to a previous publications outlining five subsets of effector and memory cells (Appay et al., 2008). By averaging the files of multiple healthy donors, the correlation of transitions in percent relative intensity of markers could be determined. The averaged modeled data of 20 healthy donors showed that down-regulation of CD45RA and CCR7 at the end of the naïve stage is significantly correlated (Fig. 4). These transitions in expression levels define the end of the naïve stage and the beginning of the CM stage. There is no evidence that later changes in CCR7 form an additional stage. The indicator for the end of the CM stage and the beginning of the EM stage is defined by the down-regulation of CD28 and the up-regulation of CD45RA.