Plant-based oral tolerance induces CD4⁺CD25⁺ Tregs and LAP⁺CD25⁻CD4⁺ Tregs. (A) Experimental outline of adoptive transfer. CD4⁻, CD4⁺CD25⁻, and CD4⁺CD25⁺ cells were purified via magnetic sorting from spleens and MLNs of FIX-fed/intravenously challenged hemophilia B mice and pooled. Doses of 10⁶ cells per mouse were adoptively transferred into strain-matched mice via tail vein injection. Control cells were from naive mice. After 24 hours, all recipient mice (n = 5-8 per group) were challenged with 1 IU FIX in adjuvant via subcutaneous injection. IgG titers against FIX (B) were determined 3 weeks later. (C) Purified CD4⁺CD25⁻ cells were further divided into LAP⁺ and LAP⁻ cells via magnetic sorting. The cells were adoptively transferred (n = 4-6 per group), followed by immunization with FIX. IgG titers against FIX (D) were determined 3 weeks later. Control mice had received cells from naive mice. Data are average ± SEM. Shown are data from 2 independent experiments that showed similar results.