Recombinant interferon alpha-2a is an active formula for the treatment of various cancer cells like malignant melanoma and a variety of virus infection diseases like acute or chronic hepatitis. The bioassay system based on the measurement of virus inhibitory activity by interferon has been used for interferon analysis with low repeatability. Here, we developed the HPLC assay to measure reproducibly interferon alpha-2a protein content which is replaceable to the reported bioassay. This method separated interferon alpha-2a from its oxidized forms and human serum albumin used as excipients. The regression coefficient using interferon alpha-2a EP CRS is more than 0.9 in the range from 5 ${mu}g/ml$ to 200 ${mu}g/ml$. The recovery result in the range from 15 ${mu}g/ml$ to 60 ${mu}g/ml$ is $97{sim}104%$ and the precision is $0.2{sim}1.7%$. The interferon alpha contents of 5 products are about 30 ${mu}g/ml$.