Mog35 55 peptide

EAE was induced by immunization female mice with MOG_35–55 peptide emulsified in CFA (Difco Laboratories) at a dose of 100µg (C57BL/6 and F1 mice) or 150µg (NOD mice) per mouse, followed by the administration of pertussis toxin (150 ng per mouse; List biological laboratories, Inc.) on days 0 and 2 as described^{14 (link)}. Clinical signs of EAE were assessed according to the following score: 0, no signs of disease; 1, loss of tone in the tail; 2, hind limb paresis; 3, hind limb paralysis; 4, tetraplegia; 5, moribund.GCV (APP Pharmaceuticals), or vehicle control (PBS) was administered daily (25mg/kg, subcutaneously) 7 days before EAE induction and continued for the duration of the acute phase (day 15), or only during the progressive/chronic phase (day 30–50). LacCer (Matreya LLC) or vehicle (10% DMSO), were administered at a dose of 10µg per mouse together with the MOG_35–55 peptide to C57BL/6J mice during EAE induction, and also intraperitoneally (i.p.) every other 3 days henceforth. LacCer or vehicle administration to NOD mice were initiated 35 days after disease induction at a dose of 10µg per mouse given i.p. every 3 other days. PDMP (Matreya LLC) or vehicle control (5% tween-80), were administered at day 40 after EAE induction twice a day with 20mg/kg given i.p. for the duration of the experiment.

Mice were immunized subcutaneously with 100 μg MOG_35–55 peptide (MEVGWYRSPFSRVVHLYRNGK) emulsified in CFA (Difco Laboratories) with 500 μg Mycobacterium tuberculosis on day 0. Mice also received 200 ng of pertussis toxin (List Biological Laboratories, Inc.) by intraperitoneal injection on days 0 and 2. Classical disease severity was assessed daily by assigning clinical scores according to the following ascending paralysis scale: 0, no disease; 1, tail paralysis; 2, weakness of hind limbs; 3, paralysis of hind limbs; 4, paralysis of hind limbs and severe hunched posture; 5, moribund or death. The incidence of atypical disease was recorded based on the development of balance issues and ataxia. To evaluate the role of IL-4 in Nlrp12-mediated atypical neuroinflammatory disease, EAE was induced in WT and Nlrp12^−/− mice as described above. Mice received either vehicle control or 500 μg anti-IL-4 blocking antibody (11B11) by intraperitoneal dosage on days -1, 0, 2, 4, 6, 8, and 13, and classical clinical scores were assigned based on ascending paralysis development.

For induction of EAE, mice were injected subcutaneously (s.c.) into the base of the tail with a volume of 100 μl containing 100 μg/ml MOG35–55 peptide (BD Biosciences) emulsified in complete Freund’s adjuvant (CFA), as we have described [16 (link), 20 (link)]. Mice were also injected intraperitoneally (i.p). with 200 ng of pertussis toxin on days 0 and 2, and clinical symptoms were scored regularly according to the following criteria: 0, no symptoms; 0.5, partially limp tail; 1, completely limp tail; 1.5, impaired righting reflex; 2, hind limb paresis; 2.5, hind-limb paralysis; 3, forelimb weakness; 4, complete paralysis; 5, death.