Intracellular delivery of proteins offers an alternative to genetic modification or siRNA transfection for functional studies of proteins in live cells, especially for studies in cancer cells for therapeutics development. However, lack of efficient and biocompatible delivery system has limited the use of protein for in vitro cancer research. In this study, we design and evaluate an amphipathic peptide RV24, composing of a hydrophobic domain for protein binding, a flexible linker, and a hydrophilic domain to facilitate cell penetration. When using beta-galactosidase as a cargo protein for comparison with commercially available peptide- and lipid-based carriers. RV24 peptide provides up to 5-fold, increase in quantity delivered into 3 different cancer cell lines. Green fluorescent protein could also be delivered rapidly within 4 h and transduced up to 83% of tested cancer cell lines. Although having a cell penetrating domain, RV24 peptide did not compromise cell viability, morphology and granularity significantly. These findings suggest the feasibility of using biocompatible amphipathic peptide to efficiently deliver protein-based molecules intra-cellularly for in vitro cancer research. (C) 2012 Elsevier Inc. All rights reserved.