Purification of Anti Spike SARS-CoV-2 Monoclonal Antibodies Using Ion Exchange Chromatography with Different pH Conditions

Tika Widayanti, Mar'atul Azizah, Retno Lestari, Jodi Suryanggono, Febby Nurdiya Ningsih, Sabar Pambudi

Research output: Contribution to journalConference articlepeer-review


The development of antibody purification is challenging since the process aims to obtain abundant yield at a low cost while maintaining the level of product purity and quality. Purification of antibodies by ion exchange chromatography is widely used in monoclonal antibodies manufacturing. Several factors influence the binding characteristics of IgG, such as pH, conductivity or salt concentration, and linear velocity of sample loading into the column. The net surface charge of immunoglobulins, consisting of many different amino acids containing weak acidic and basic groups, will change gradually as the pH of the solution changes. In this study, we performed a simple method of anion-exchange chromatography to selectively remove impurities, using elution buffers with three different pH. Diethylaminoethyl (DEAE) sepharose fast-flow columns were used as weak anion exchanger to obtain high purity levels for IgG antibodies with a broad range of isoelectric points (pI). Increasing pH during anion-exchange chromatography increases the concentration of the dissociated form of weak acidic groups as eluant. SDS-PAGE gel electrophoresis was used to visualize the protein profiles, and monoclonal antibodies activity was determined with indirect ELISA. The result showed that anion-exchange DEAE resins were able to remove protein impurities compared to unpurified mixtures. DEAE column with Tris-Cl buffer pH 8.0 provided optimal conditions compared to pH 8.5 and 9.0. We employed a gradient salt elution procedure and achieved a high peak with 2ml fractions from 250ul of BALB/c mice ascites fluid. However, additional steps for the downstream antibody purification process are needed to meet the industrial standards.

Original languageEnglish
Article number070003
JournalAIP Conference Proceedings
Issue number1
Publication statusPublished - 7 Mar 2024
Event15th Asian Congress on Biotechnology in conjunction with the 7th International Symposium on Biomedical Engineering, ACB-ISBE 2022 - Bali, Indonesia
Duration: 2 Oct 20226 Oct 2022


  • antibody purification
  • ion-exchange chromatography
  • monoclonal antibodies


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