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文章利用仿生多肽与腺相关病毒(AAV)的亲和相互作用，将仿生多肽偶联于琼脂糖微球，制备出一种用于AAV纯化的通用型亲和介质。以不同种类的仿生多肽为配基，以Sengarose 4FF微球为基质，通过环氧活化的方式将多肽配基偶联至琼脂糖微球，制备AAV多肽亲和介质；通过调控活化基团密度、多肽配基种类及浓度、反应温度等条件，最终制备出配基密度为2.2～5.6 mg/mL的多种AAV多肽亲和介质。该介质形貌规整，平均粒径为90μm。扫描电子显微镜结果表明，琼脂糖微球在偶联配基前后表面形貌基本未改变。在介质吸附性能研究中，分别采用高效液相色谱(HPLC)和激光共聚焦显微镜(CLSM)表征介质结合AAV的效果。结果表明，该仿生多肽介质能有效结合AAV，在特定吸附与解吸条件下顺利捕获目标病毒。进一步研究该多肽亲和介质的通用结合性能发现，其对AAV2型和AAV9型均具有结合能力。多肽配基密度对AAV2吸附量的影响结果表明，介质对AAV的结合量随配基密度增大而增加，当配基密度为5.6 mg/mL时，AAV结合量达到最大值。文章为AAV亲和层析纯化技术的发展提供了新思路。
Abstract：The article utilized the affinity interaction between biomimetic peptides and adeno-associated virus(AAV) to couple the biomimetic peptides onto agarose microspheres, thereby preparing a universal affinity medium for AAV purification. With different types of biomimetic peptides as ligands and Sengarose 4 FF microspheres as the matrix, the biomimetic peptide ligands were coupled to the agarose microspheres through epoxy activation to prepare AAV peptide affinity media. By regulating the density of the activated groups, the type and concentration of the peptide ligands, and the reaction temperature, various AAV peptide affinity media with ligand densities ranging from 2.2 to 5.6 mg/m L were ultimately prepared. The morphology of the medium was regular, with an average particle size of 90 μm. Scanning electron microscopy results indicated that the surface morphology of the agarose microspheres remained largely unchanged before and after coupling the ligands. In the study of the adsorption performance of the medium, high-performance liquid chromatography(HPLC) and confocal laser scanning microscopy(CLSM) were used to characterize the binding effect of the medium on AAV. The results showed that the biomimetic peptide medium could effectively bind AAV and successfully capture the target virus under specific adsorption and desorption conditions. Further research on the universal binding performance of the peptide affinity medium revealed that it had binding capabilities for both AAV2 and AAV9 types. The results of the influence of the peptide ligand density on the adsorption capacity of AAV2 indicated that the binding capacity of the medium to AAV increased with the increase in ligand density, reaching the maximum value whenthe ligand density was 5.6 mg/mL. The article provided a new idea for the development of AAV affinity chromatography purification technology.
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基本信息:
DOI：10.16026/j.cnki.iea.2025060455
中图分类号:Q503;O647.3
引用信息:
[1]薛瑞雪,黄永东,赵岚,等.腺相关病毒通用型亲和介质的制备与吸附性能研究[J].离子交换与吸附,2025,41(06):455-463.DOI:10.16026/j.cnki.iea.2025060455.
基金信息:
国家重点研发计划资助项目(项目号2023YFC2812001,2023YFC2812002); 国家自然科学基金资助项目(项目号22278411,32471385)