E-peptide of the pro-Insulin-like growth factor-I (pro-IGF-I) is produced from pre-pro-IGF-I by proteolytic cleavage in the post-translational processing. The human Eb-peptide (hEb-peptide), derived from the E domain of pro-IGF-IB isoform, is a bioactive molecule whose exact physiological role remains elusive. Accumulated evidence reported from our laboratory indicated that hEb-peptide possesses activity against multiple hallmark characteristics of solid tumor in different cancer cell types. In human breast carcinoma cells (MDA-MB-231), it was demonstrated that hEb-peptide can promote cell attachment to substratum, inhibit colony formation in a semisolid medium, reduce cancer cell invasion, and inhibit cancer-induced angiogenesis. Like the action of other peptide hormones, these cellular responses triggered by hEb may be initiated through binding to a receptor molecule residing on the surface of the cell. Our laboratory and the others have previously provided evidence demonstrating the existence of hEb-peptide specific binding components residing on the cell membrane. In this study, we report the isolation and identification of eight protein molecules bound reversibly with hEb-peptide from the membrane preparation of MDA-MB-231 cells. Some of the identified proteins are known to be present at cell surface and function as receptors while the others are not. The functions of these molecules reveal strong correlation with the demonstrated activities of hEb-peptide on MDA-MB-231cells, suggesting hEb-peptide activity on cancer cells might be mediated by these molecules.