Recombinant interferon-alpha (rIFN-alpha) has been widely used for treating viral infections. However, the clinical efficacy of unmodified rIFN-alpha is limited due to small molecular size and rapid clearance from circulation. In this study we developed a novel strategy for half-life extension of porcine IFN-alpha (PoIFN-alpha) by fusion to the immunoglobulin (Ig)-binding C2 domain of streptococcal protein G (SPG). The coding sequences for PoIFN-alpha 6 and SPG C2 domain, with a tobacco etch virus (TEV) protease recognition sequence introduced at the 5-end, were cloned into an elastin-like polypeptide (ELP) fusion expression vector and expressed as an ELP-PoIFN alpha-C2 fusion protein. After optimization of the conditions for soluble protein expression and purification, the fusion protein was purified to more than 90% purity by two rounds of inverse transition cycling (ITC) in the presence of 0.5% Triton X-100. After cleavage with self-aggregating peptide ELK-16-tagged tobacco etch virus protease, the protease was removed by quick centrifugation and PoIFN alpha-C2 protein was recovered by an additional round of ITC with 98% purity. Western blotting analysis showed that PoIFN alpha-C2 protein had the specific affinity for pig IgG binding. The antiviral assay showed that PoIFN alpha-C2 protein had potent antiviral activities against vesicular stomatitis virus and porcine pseudorabies virus. After single intravenous or subcutaneous injection into rats, PoIFN alpha-C2 protein showed 16- or 4-fold increase in serum half-life with significantly improved bioavailability.