Based on the hypothesis that interactions between virions and serum components may influence the outcome of dengue virus (DENV) infections, we decided to use affinity chromatography with domain III from the envelope (E) protein of DENV2 (DIIIE2) as a ligand to isolate virus-binding proteins from human plasma. This approach yielded serum amyloid P (SAP) and α₂-macroglobulin (α₂M) as novel viral interactors. After confirming the specific binding of both SAP and α₂M to DIIIE2 by ELISA, the latter interaction was examined in greater detail. We obtain evidence suggesting that the binding species was actually the receptor-activated form of α₂M (α₂M*), that α₂M* could bind monovalently to recombinant domain III from all four DENV serotypes with affinities in the micromolar range ranking as DENV4>DENV1~DENV2>DENV3 and that this interaction exhibited a strong avidity effect when multivalent binding was favoured (K_D8×10⁻⁸M for DIIIE2). We also showed that α₂M* bound to DENV virions of the four serotypes, protecting the virus from temperature-induced inactivation in the absence of serum and enhancing infectivity. The latter effect exhibited an ED₅₀of 2.9×10⁻⁸M, also suggesting an avidity effect due to multivalent binding. These results will further contribute to the characterization of the virus–host factor interaction network during human DENV infection.