Thymosin alpha 1-thymopentin (T alpha 1-TP5) fusion peptide has been proved to be an immune regulator based on its higher immunoregulatory activity than T alpha 1 and TP5. To obtain T alpha 1-TP5 more effectively and economically, T alpha 1-TP5 was genetically fused to a self-cleaving intein-chitin binding domain tag for purification via chitin beads in Escherichia coli. After affinity purification, the target peptide was released from the chitin beads via self-cleaving intein ((INTervening protEIN) induced by dithiothreitol. Further, T alpha 1-TP5 was purified by Superdex 30 and identified by Tricine-SDS-PAGE and electrospray ionization-mass spectrometry. Finally, about 7.6 mg T alpha 1-TP5 purified from the soluble fraction and inclusion bodies was obtained from 1 L culture media. The purity was 95% after a series of chromatographic purification steps. In vitro, the purified T alpha 1-TP5 could stimulate the proliferation of mouse splenic lymphocytes. Overall, this work demonstrated that T alpha 1-TP5 was purified with low cost and high efficiency, greatly expanding its potential use as an immune regulator. (c) 2012 Elsevier Inc. All rights reserved.