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Purification of flavivirus VLPs by a two-step chomatographic process

机译：双层缩细过程纯化了黄病毒VLP

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Flaviviruses are enveloped viruses with positive-sense, single-stranded RNA, which are most commonly transmitted by infected mosquitoes. Zika virus (ZIKV) and yellow fever virus (YFV) are flaviviruses that have caused significant outbreaks in the last few years. Since there is no approved vaccine against ZIKV, and since the existing YF attenuated vaccine presents disadvantages related to limited supply and to rare, but fatal adverse effects, there is an urgent need for new vaccines to control these diseases. Virus-like particles (VLPs) represent a recombinant platform to produce safe and immunogenic vaccines. Thus, based on our experience of expressing in recombinant mammalian cells VLPs of most flaviviruses circulating in the Americas, this work focused on the evaluation of chromatographic purification processes for zika and yellow-fever VLPs. The clarified cell culture supernatant was processed by a membrane-based anion-exchange chromatography and then a multimodal chromatographic step. With this process, it was possible to obtain the purified VLPs with a yield (including the clarification step) of 66.4% for zika and 68.1% for yellow fever. DNA clearance was in the range of 99.8-99.9%, providing VLP preparations that meet the WHO limit for this critical contaminant. Correct size and morphology of the purified VLPs were confirmed by dynamic light scattering (DLS) and transmission electron microscopy (TEM). The promising results obtained for both zika and yellow fever VLPs indicate that this process could be potentially applied also to VLPs of other flaviviruses. (C) 2019 Elsevier Ltd.
机译：黄病毒是患有阳性意义，单链RNA的包膜病毒，这些病毒最常被感染的蚊子传播。 Zika病毒（Zikv）和黄热病病毒（YFV）是在过去几年中引起重大爆发的黄病毒。由于没有针对ZIKV的批准疫苗，并且由于现有的YF减毒疫苗具有有关供应有限的缺点，并且罕见，但致命不良反应，迫切需要新的疫苗来控制这些疾病。病毒样颗粒（VLP）代表重组平台以产生安全和免疫原性疫苗。因此，根据我们在美洲循环中循环的大多数黄病毒的重组哺乳动物细胞VLP中表达的经验，这项工作主要集中在ZIKA和黄热vlps的色谱纯化过程的评价。通过基于膜的阴离子 - 交换色谱法加工澄清的细胞培养上清液，然后加工了多数制色谱步骤。通过该方法，可以获得Zika的产率（包括澄清步骤）的纯化的VLP，为黄热发热68.1％。 DNA清除的范围为99.8-99.9％，提供符合该关键污染物的人限制的VLP制剂。通过动态光散射（DLS）和透射电子显微镜（TEM）来确认纯化VLP的正确尺寸和形态。 Zika和黄热病VLP获得的有希望的结果表明，该过程可能也可能应用于其他黄病毒的VLP。 （c）2019年elestvier有限公司

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