Optimization of conditions for single-step purification of recombinant hepatitis B surface antigen produced in Pichia pastoris using ion exchange chromatography |
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| Authors: | Nazanin Ashourian Moghadam Farshid Pajoum Shariati Hooman Kaghazian |
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| Affiliation: | 1. Department of Chemical Engineering, Islamic Azad University, Science and Research Branch, Tehran, Iran;2. Production and Research Complex, Pasteur Institute of Iran, Karaj, Iran |
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| Abstract: | ABSTRACTThe adsorption and release of rHBsAg extracted from the final dosage form on various ion exchange resins and under different pH conditions were investigated after its peptide map and isoelectric point (PI) determination. Efficient antigen adsorption to the anion exchange resins occurred when the pH value of the protein buffer was adjusted to 5.0. In purification of rHBsAg derived from the yeast crude extract using Q Sepharose FF column, with adjusting the pH value of the crude extract to 5.0 (i.e., near to the target protein PI) and using 2M NaCl, rHBsAg with high purity (up to >95%) was obtained.Abbreviations: rHBsAg, recombinant hepatitis B surface antigen; Alhydrogel, aluminum hydroxide; IEF, isoelectric focusing; PI, isoelectric point; SDS-PAGE, sodium dodecyl sulfate-polyacrylamide gel electrophoresis; RP-HPLC, reversed-phase high-performance liquid chromatography; SE-HPLC, size-exclusion high-performance liquid chromatography; PBS, phosphate-buffered saline |
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| Keywords: | Recombinant hepatitis B surface antigen (rHBsAg) separation purification ion exchange chromatography |
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