Mog35 55 peptide

EAE was initiated using a myosin oligodendrocyte protein (MOG)/Bordetella pertussis toxin (PT) method [15 (link)]. Briefly, mice were subcutaneously injected with 0.2 μmol of MOG_35-55 peptide (MEVGWYRSPFSRVVHLYRNGK, synthesized at the University of Utah HSC Core) in complete Freund’s adjuvant (CFA, Sigma, 2 mg/mL). Two hundred nanograms of PT (Sigma) was injected into the mice twice intravenously. Clinical scores were determined based on the following criteria: 0, no clinical disease; 1, loss of tail tonicity; 2, mild hind limb paresis; 3, moderate hind limb paralysis; 4, paraplegia; 5, quadriplegia, coma, or death. For tissue analysis, animals were sacrificed at peak disease (days 20–21).

EAE was induced in 10- to 12-week-old, female C57Bl/6 mice by subcutaneous injection of 100 μg MOG_35–55 peptide emulsified in complete Freund’s adjuvant (Sigma-Aldrich) supplemented with 400 μg Mycobacterium tuberculosis (Difco) into both hind limb flanks. Mice received an intraperitoneal injection of 350 ng Pertussis toxin on day 0 and day 2. Mice were monitored daily and scored blind to avoid unconscious bias. Clinical scores were assigned according to an arbitrary clinical scale: 0, no detectable impairment; 0.5, partial loss of tail tone; 1, complete loss of tail tone; 1.5, complete loss of tail tone and difficulty righting; 2, complete loss of tail tone and weak hind limbs; 2.5, one paralysed hind limb; 3, complete paralysis of hind limbs; 3.5, complete paralysis of hind limbs and ascending paresis affecting the trunk region; 4, complete paralysis of hind limbs and paresis in forelimbs; and 5, moribund or deceased. Under recommendation of the animal ethics committee, mice were euthanised upon reaching a clinical score of 3.5. In some experiments, mice were euthanised on Day 9. Blood was collected via cardiac bleed into heparin-coated tubes, red cell lysed and processed for flow cytometric analysis.