A Preparative Purification Process for Recombinant Hepatitis B Core Antigen Using Online Capture by Expanded Bed Adsorption Followed by Size-Exclusion Chromatography

Abstract

Hepatitis B core antigen (HBcAg) is an important serological marker used in the diagnosis of hepatitis B virus (HBV) infections. In the current study, a fast and efficient preparative purification protocol for truncated HBcAg from Escherichia coli disruptate was developed. The recombinant HBcAg was first captured by anion exchange expanded bed adsorption chromatography integrated with a cell disruption process. This online capture process has shortened the process time and eliminated the "hold-up" period that may be detrimental to the quality of target protein. The eluted product from the expanded bed adsorption chromatography was subsequently purified using size-exclusion chromatography. The results showed that this novel purification protocol achieved a recovery yield of 45.1% with a product purity of 88.2%, which corresponds to a purification factor of 4.5. The recovered HBcAg is still biologically active as shown by ELISA test.

Purification of histidine-tagged nucleocapsid protein of Nipah virus using immobilized metal affinity chromatography

“…As a summary, the purification of his-tagged N protein of NiV using IMAC packed bed purification process has achieved a 68.3% yield and a purification factor of 7.94. Further improve of the histidine-tagged N protein purity can be achieved with size exclusion chromatography as been described by Ho et al [29]. The purified N protein of NiV from the IMAC system was evaluated by ELISA using the anti-N protein rabbit serum.…”

Recovery and purification of recombinant 503 antigen of Leishmania infantum chagasi using expanded bed adsorption chromatography

“…Ho et al [42] reported that the use of an expanded bed allows the combination of many of the conventional primary recovery steps, such as clarification, concentration, and initial purification, into a single process. Furthermore, a typical cycle of purification by EBA chromatography including equilibration (30 min), loading (30 min), washing (60 min) and elution (30 min) only took 2.5-3.5 h, making the process attractive for large-scale use [43].…”

Gel-Filtration Chromatography

“…Size-exclusion chromatography was used downstream of expanded bed adsorption chromatography to recover active recombinant hepatitis B core antigen (HbcAg) in 45 % yield with a purifi cation factor of 4.5 [ 18 ]. A Sephacryl S-1000 SF proved to be effective and economical in the purifi cation of recombinant Bombyx mori nucleopolyhedrosis virus displaying human pro renin receptor [ 19 ].…”