Phenotypic analysis of peripheral B-cell subpopulations in patients with ITP. Peripheral blood from patients and healthy controls were stained with CD19, CD24, CD38, and CD27. Patients listed in Tables 1 through 3 were analyzed: nonsplenectomized ITP cohort without treatment with low platelet counts, nonsplenectomized group on treatment with TPO agents with increased platelet counts, splenectomized patients with platelet counts of less than 50 × 10⁹/L without ITP treatment, and splenectomized patients on treatment with TPO agents with platelet counts of more than 50 × 10⁹/L compared with healthy controls (n = 13; median age, 40 years; range, 24-66; 8 males and 5 females). (A) CD24 and CD38 expression patterns of CD19⁺ lymphocyte gated cells are shown in the representative dot plots of healthy controls and ITP patients. Three distinct subpopulations have previously been described in human peripheral blood³ : CD19⁺CD24^hiCD38^hi cells that include immature B cells, CD19⁺CD24^intCD38^int consisting primarily of mature B cells and CD19⁺CD24⁺CD38⁻ with mostly memory B cells. The gating strategy for analysis of the CD24^hiCD38^hi B cells is indicated. In supplemental Figure 2, the gating strategy for the other 2 subpopulations are shown. (B) Frequency of CD24^hiCD38^hi of CD19⁺ subset in healthy controls and nonsplenectomized or splenectomized ITP patients with platelet counts of less or more than 50 × 10⁹/L is indicated. P values shown highlight the statistically significant reduction in CD19⁺CD24^hiCD38^hi subpopulation frequency in nonsplenectomized ITP patients with platelet counts less than 50 × 10⁹/L compared with healthy controls, and also indicate a significant increase in this B-cell population in patients on treatment with TPO agents whose platelet counts are above 50 × 10⁹/L. Although the P value is not shown, splenectomized patients with platelet counts less than 50 × 10⁹/L have increased frequency of CD19⁺CD24^hiCD38^hi compared with nonsplenectomized patients off treatment with low platelet counts (P = .02). (C) Representative dot plots of CD19 and CD27 memory surface marker expression pattern in healthy controls and patients are shown with gating strategy used to determine the proportion of CD27⁺ cells in B cells. (D) Frequency of CD19⁺CD27⁺ in the controls and patient groups in panel B are shown in various cohorts. *Statistically significant differences were observed in splenectomized patients regardless of platelet counts compared with controls.