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Amyloid beta-protein (Aβ), in its soluble form, is known to bind several circulatory proteins such as apolipoprotein (apo) E, apo J and transthyretin. However, the binding of Aβ to intracellular proteins has not been studied. We have developed an overlay assay to study Aβ binding to intracellular brain proteins. The supernatants from both rat and human brains were found to contain several proteins that bind to Aβ 1–40 and Aβ 1–42. No major difference was observed in the Aβ binding-proteins from brain supernatants of patients with Alzheimer's disease and normal age-matched controls. Binding studies using shorter amyloid beta-peptides and competitive overlay assays showed that the binding site of Aβ to brain proteins resides between 12–28 amino acid sequence of Aβ. The presence of several intracellular Aβ-binding (AβB) proteins suggests that these proteins may either protect Aβ from its fibrillization or alternatively promote Aβ polymerization. Identification of these proteins and their binding affinities for Aβ are needed to assess their potential role in the pathogenesis of Alzheimer's disease.