Experimental Immune Thrombocytopenia Mouse Model

IVIg (Gamimune N) was purchased from Bayer (Whippany, NJ, USA). Experimental ITP was induced as described previously [18 (link),19 (link),82 (link),83 (link)]. The mice were intravenously injected with 0.1 mg/kg (body weight) of an antiplatelet monoclonal antibody (rat antimouse integrin α_IIb/CD41 Ig, clone MWReg30, BD Biosciences, Franklin Lakes, NJ, USA) to induce ITP. To analyze the platelet count, whole blood samples (100–120 μL) of the mice were collected with an anticoagulant, a citrate dextrose solution (38 mM citric acid, 75 mM sodium citrate, and 100 mM dextrose), in Eppendorf tubes. Subsequently, platelet counts were measured using a hematology analyzer (KX-21N, Sysmex, Kobe, Japan) at various time intervals [77 (link),78 (link),84 (link)]. To investigate the ameliorative effects of intravenous treatment with IVIg (high dose, 2 g/kg; vehicle: saline; injection at 0 h), cold (4 °C) exposure and restraint stress for 9 h was implemented. Anti-CD41 Ig was treated together with these treatments (IVIg, cold, restraint stress).