Collagen-induced arthritis (CIA) is an autoimmune arthritis that can be elicited by the immunization of genetically susceptible strains of mice with type II collagen (CII). We have analysed the molecular interactions that occur between an arthritogenic T-cell determinant CII (442-457) and the murine class II susceptibility allele I-A(r). To determine which amino acid residues within the CII (442-457) sequence are responsible for binding to I-A(r), a soluble I-A(r):IgG2aFc fusion protein-peptide binding assay was developed. Various concentrations of analogue peptides were tested for their ability to compete with biotinylated CII (607-622) for binding to I-A(r), thereby establishing a relative comparison of the binding affinities among these analogues. Analogue peptides with substitutions at positions 447 (Ala —> Val), 448 (Gly —> Ala) and 451 (Gly —> Ala) bound poorly to the I-A(r) molecule. These data suggest that positions 447, 448 and 451 on CII are the major anchor points to I-A(r) molecules. In cytokine assays, only substitutions within positions 445-454 decreased the interferon-gamma production by T cells. These data narrow the core of the arthritogenic T-cell determinant to CII (445-454). Identification of the molecular interactions involved in T-cell recognition of CII should lead to antigen-specific means of inhibiting autoimmune arthritis.