Up to 90% of circulating insulin-like growth factors (IGF-I and IGF-II) are carried in heterotrimeric complexes with a binding protein (IGFBP) and a liver-derived glycoprotein known as the acid-labile subunit. IGFBP-3 is considered unique among the six well characterized IGFBPs in its ability to complex with the acid-labile subunit. However, a basic carboxyl-terminal domain of IGFBP-3, known to be involved in its interaction with the acid-labile subunit, is shared by IGFBP-5, suggesting the possibility of ternary complexes containing IGFBP-5. We now demonstrate using three independent methods that human IGFBP-5, when occupied by IGF-I or IGF-II, forms ternary complexes of approximately 130 kDa with the acid-labile subunit. IGFBP-3 competes with approximately twice the potency of IGFBP-5 for the formation of such complexes. No other IGFBP complexes with the acid-labile subunit itself or competes with IGFBP-5 for complex formation. As observed for IGFBP-3, ternary complexes containing IGFBP-5 form preferentially in the presence of IGF-I, even though IGFBP-5 has a preferential affinity for IGF-II over IGF-I. By size fractionation chromatography, serum IGFBP-5 co-elutes predominantly with ternary complexes. The demonstration of IGFBP-5-containing ternary complexes indicates an unrecognized form of IGF transport in the circulation and an additional mechanism for regulating IGF bioavailability.