SP Focurose HPR Purification Process for Porcine Circovirus Type 2 (PCV-2) Cap Protein Antigen Expression
Suzhou Vdo Biotech Co.,Ltd
Suzhou Vdo Biotech Co., Ltd. is a specializes in production of biotechnology microspheres.
Porcine Circovirus Type 2 (PCV-2) is a pathogenic virus and the primary causative agent of post-weaning multisystemic wasting syndrome in piglets. The capsid protein (Cap protein) of PCV-2 is its major structural protein and the primary immunogenic protein (i.e., antigen). The effectiveness of vaccines also relies on this protein. The level of stress response in pig herds is directly proportional to the impurities present in the vaccine. Therefore, the separation of PCV-2 Cap protein from various impurities such as miscellaneous proteins, liposomes, nucleic acids, pigments, and endotoxins poses a challenge and a technical focal point.
Next, we will introduce the purification scheme for Porcine Circovirus (PCV) Cap Protein.
The SP Focurose HPR produced by VDO is a cation exchange medium prepared using highly cross-linked agarose microbeads as the matrix and connected with sulfopropyl ligands. It has an average particle size of 70±5μm and allows the separation of PCV Cap protein antigen and impurities based on their different charges in the buffer solution with a pH range of 6.0-7.0.
The specific steps of the chromatographic method are as follows:
The collected supernatant from SF9 insect cells is centrifuged at 25°C and 11,500 rpm for 30 minutes. The supernatant is then filtered through a 0. xn--45m-yyc membrane and adjusted to a pH of 6.5-7.0 using 0.2M HCl, resulting in the purified sample solution.
The SP Focurose HPR cation exchange medium is packed into a chromatography column of a certain specification. The column efficiency should be above 3600 plates per meter (PPM), and the symmetry factor should be between 0.8 and 1.8 to ensure normal column usage.
Prepare a pH 6.5-7.0, 20-50mM phosphate buffer solution as the equilibration and wash buffer and a pH 6.5-7.0, 20-50mM phosphate buffer solution containing 1M NaCl as the elution buffer.
The flow rate throughout the chromatographic process should be between 60-120 cm/h. The optimal flow rate is selected based on actual conditions such as column pressure difference and retention time, ensuring that the equilibrium buffer is passed through the column at 3-10 times the column volume until its pH and conductivity values stabilize and the UV280 reading is zeroed. Then, 10-20 times the column volume of the purified sample solution is loaded for the sample application. After sample loading, several column volumes of the equilibration buffer are used for washing until the pH and conductivity values stabilize, and the UV280 reading is below 50 mAU. The washing volume may vary depending on the column size and sample type. Finally, elution is performed using 5-10 times the column volume of the elution buffer, and the eluted peak is collected based on the UV280 reading. The collected eluate is the high-purity PCV Cap protein antigen.
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VDOBIOTECH Company Profile
VDO Biotech Co., Ltd. is an esteemed high-tech enterprise comprising a workforce of over 500 employees, dedicated to innovative microsphere technologies and the production of a variety of premium microsphere products catering to discerning customers worldwide. VDO is headquartered in the BioBAY of Suzhou Industrial Park in, China, with manufacturing facilities certified to ISO9001:2015 and covering more than 10,000 square meters. For any microsphere-related applications, our reliable team is the ideal partner.
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Telephone：+86-15850246572
Email:[email protected]
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