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4 weeks after the last injection of OVA-CFA, the knee joints were injected with 1 ml of OVA once a week.
Beginning five days after the second injection, 1 ml of OVA (5 mg/ml in 0.9% NaCl) was injected into the knee joints on a weekly basis over the following four weeks.
Five days after the second injection, 1 ml of OVA (5 mg/ml in 0.9% NaCl) was injected intra-articularly into the knee joint on a weekly basis over the following four weeks.
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Briefly, to a mixture of 90 mg of liposomes and 6 mg of OVA in 2.5 ml phosphate buffer (pH 7.2), 0.5 ml of 2.5% glutaraldehyde solution was added in dropwise fashion.
Briefly, mice were injected subcutaneously (s.c). on days 1 and 7 with 0.2 ml of a solution containing 100 mg of OVA and 70 mg of aluminum hydroxide.
Lymphocytes (5×106) were incubated with 2 µM of OVA (257-264) peptine in the presence of 2 µg/ml of anti-CD28 mAb (clone 37.51) for 6 hr.
A 2-ml suspension of DSS-introduced liposome and 0.5 ml of 5-mg/ml OVA peptide solution were mixed and stirred for 3 days at 4°C.
Briefly, animals were given two intradermal injections of 1 ml ovalbumin (OVA) (4 mg/ml; Sigma-Aldrich, St . Louis MO, USA) in Freund's complete adjuvant (Difco, Detroit, MI, USA).
Acute monoarticular arthritis was induced in both knee joints two weeks after the booster shot by intra-articular injection of OVA dissolved in 0.5 ml saline.
For the second and third sensitizations, patches were soaked in 100 ml of PBS containing 100 mg OVA and 50 μg of the synthetic corticosteroid triamcinolone acetonide or 50 μg of the test drugs.
Forty-eight hours later each mouse received an injection of 0.1 ml OVA (1 mg/ml) containing 4% Evans blue via the tail vein.
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