The present invention provides a single domain antibody (sdAb) scaffold comprising one or more than one non-canonical disulfide bond in the framework region (FR). The one or more than one non-canonical disulfide bond may be formed between cysteines introduced by mutations in FR2 and FR3. In the case where the sdAb scaffold is a VH, the Cys may be introduced at any one of positions 47-49 and any one of positions 67-71, based on Kabat numbering； in one example, the Cys may be introduced at positions 49 and 69, based on Kabat numbering. In the case where the sdAb scaffold is a VL, the Cys residues may be introduced at any one of positions 46-49 and any one of positions 62-66, based on Kabat numbering； in one example, the Cys residues may be introduced at positions 48 and 64, based on Kabat numbering.
