Fig. 1.

Gating strategy for six-colour flow cytometric detection of the total CD4⁺CD127^low+/−CD25⁺forkhead box protein 3 (FoxP3⁺) regulatory T cell (T_reg) cell pool and its percentage of naive DR^-CD45RA⁺ T_regs, DR^-CD45RA^- T_regs, DR^low+CD45RA^- T_regs and DR^high+CD45RA^- T_regs. (a) First, CD4⁺ T cells (P1) were gated by fluorescence intensity of CD4 versus side light-scatter (SSC). (b) CD4⁺CD127^low+/−CD25⁺ T_reg cells were gated by fluorescence intensity of CD25 versus CD127 (P2). (c) CD4⁺CD127^low+/−CD25⁺FoxP3⁺ T_reg cells were gated by excluding cells without FoxP3 expression (P3). (d) The percentage of the DR^high+CD45RA^- (P4), the DR^low+CD45RA^- (P5), the DR^-CD45RA^- (P6) and the naive DR^-CD45RA⁺ T_reg subset (P7) was estimated by analysing CD4⁺CD127^low+/−CD25⁺FoxP3⁺ T_reg cells (P3) for their expression of human leucocyte antigen D-related (HLA-DR) and CD45RA. (a–d) Shows a representative experiment for healthy pregnancies, (e–h) show representative experiments for patients with HELLP syndrome (e), pre-eclampsia (f), CI (g) and PL (h). CI: cervical insufficiency; PL: preterm labour necessitating preterm delivery; HELLP syndrome: haemolysis-elevated liver enzyme level-low platelet count syndrome.