Chinese Hamster Ovary (CHO) cells are frequently used to produce recombinant HIV envelope protein gp120. In this study, we characterized the CHO proteins that become co-purified with gp120 when lectin affinity chromatography is used. Many of co-purified CHO proteins identified were extracellular matrix components. We verified a method for separating gp120 from these CHO proteins with anion exchange chromatography, and assessed the biochemical activity of gp120 preparations with and without co-purified CHO proteins.