Glycosylation and RNase activity are essential for omega-1 to prime Th2 responses in vivo. 4get/KN2 IL-4 dual reporter mice were injected s.c. with 20 µg SEA or 3 µg WT or mutant recombinant omega-1 into the footpad. After 7 d, the frequency of GFP⁺ and huCD2⁺ within the CD4⁺CD44^high effector T cell population was determined by flow cytometry in the draining popliteal LNs. (A–C) Depicted are concatenated FACS plots (A), frequencies of huCD2⁺ within the CD4⁺CD44^high population (B), and total huCD2⁺ T cell numbers in draining LNs (C) of combined data of four mice per group. (A) The frequencies of each population are indicated as percentages in the plots. One of three independent experiments is shown. Bars represent mean ± SD. * and #, P < 0.05; ** and ##, P < 0.01; ***, P < 0.001 for values significantly different from the PBS control (*) or between test conditions (#) based on two-sided t test. H58F, RNase mutant; N71/176Q, glycosylation mutant.