Association of the invariant chain (Ii) with MHC class II α and β chains is central for their functionality and involves the Ii CLIP(81–104) region. Ii mutants with an antigenic peptide sequence in place of the CLIP region are shown to form αβIi complexes resistant to dissociation by SDS at 25°C. This reflects class II peptide binding site occupancy, since substitution of the major anchor residue within the antigenic peptide sequence of one of these Ii mutants abolishes its capacity to form SDS‐stable heterotrimers. Therefore, CLIP location within Ii is compatible with CLIP access to the class II binding groove. However, in wild‐type Ii this access does not lead to a tight association, which seems to be affected by the Ii 81–90 region. This region, together with a region C‐terminal of CLIP, is shown to contribute to Ii association with HLA‐DR1 molecules. Thus, Ii mutants with non‐HLA‐DR1 binding sequences in place of the CLIP(87–102) region can still associate with HLA‐DR1 molecules and inhibit peptide binding.