The tumor necrosis factor-alpha (TNF alpha) plays an important role in a number of chronic inflammatory disorders. Monoclonal camelidae variable heavy chain domain-only antibodies (VHH) have been developed to antagonize the action of human TNF alpha (anti-TNF-VHH). Here we describe a strategy to obtain functional dimeric anti-TNF-VHH molecules, based on the C-terminal fusion of a kappa light chain domain to the anti-TNF-VHH. The resulting fusion protein was transiently expressed by use of viral vectors in Nicotiona benthandana((Nb)) leaves and purified. Competitive ELISA and cell cytotoxicity assays revealed that the dimerized anti-(TNF)-T-Nb-VHHC kappa proteins blocked TNF alpha-activity more effectively than either the monomeric Escherichia coli((Ec)) produced anti-(TNF)-T-Ec-VHH or the monomeric anti-(TNF)-T-Nb-VHHC kappa. We suggest that enhanced inhibition shown by dimeric anti-(TNF)-T-Nb-VHHC kappa, proteins is achieved by an increase in avidity. Biotechnol. Bioeng. 2010;106: 161-166. (C) 2009 Wiley Periodicals, Inc.