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[Purification of recombinant erythropoietin by reversed-phase high performance liquid chromatography (RP-HPLC)].

Zongshu Zou, K Sun, Lou D, Min Hu

Abstract

Recombinant erythropoietin is a glycoprotein which is strongly hydrophobic. We purified recombinant erythropoietin by RP-HPLC. The results showed that 30 nm Spherisorb C4 reversed-phase column and acetonitrile-TFA mobile phase with gradient elution, could readily isolate recombinant erythropoietin from crude samples with high speed and efficiency. Erythropoietin could be eluted at the concentration of about 50% acetonitrile with appropriate sample purity and elution gradient. When RP-HPLC is used as a final step of purification, the purity of sample should be at least 60%. The purity of the product tested by SDS-PAGE was nearly 100%. And the specific activity of the product was about 1.96 x 10(3) IU/g protein.

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Key concepts: Chromatography, High-performance liquid chromatography, Erythropoietin, Elution, Recombinant DNA, Chemistry, Acetonitrile, Gradient elution