*5.3. Induction of the EAE*

The EAE model was induced as previously described [98]. Animals were anesthetized (isoflurane 1.5–2% in oxygen) and immunized subcutaneously at the base of the tail (s.c.) with 200 μg of myelin oligodendrocyte glycoprotein (MOG35–55) peptide (Proteimax Technology, Sao Paulo, SP, Brazil) emulsified in incomplete Freund's adjuvant (IFA) supplemented with *Mycobacterium tuberculosis* (4 mg/mL, H37Ra, Difco™, Detroit, MI, USA). Intraperitoneal (i.p.) injection of 300 ng of pertussis toxin (Sigma-Aldrich™, St. Louis, MO, USA) was performed immediately after immunization and 48 h later. Control animals were treated with the vehicle (complete Freund's adjuvant—CFA, s.c. and pertussis toxin, i.p.). Animals were daily monitored according to the following clinical scores: 0.0, no symptoms; 0.5, some loss of tail tone; 1.0, loss of tail tone; 1.5, hip weakness upon ambulation; 2.0, hip weakness and partial hind limb paresis; 2.5, partial hind limb paralysis; 3.0, total hind limb paralysis but still moving on the cage; 3.5, complete hind limb paralysis and forelimbs weakness; 4.0, total hind limb paralysis and partial forelimbs paralysis; 4.5, complete hind limb paralysis, partial paralysis of the forelimbs, decreased responsiveness (consider euthanasia); 5, Immobile and unresponsive, moribund (immediate euthanasia). In the occurrence of score 3, food and water were turned available inside the cage. In the occurrence of score 4, animals were evaluated twice a day and if this score persists for three consecutive assessments, euthanasia was applied and the score of 5 was considered to the end of the period of evaluation. Body weight was measured during the course of the disease.
