Dengue fever is one of the most wide-spread vector-borne diseases in the world. Although dengue-associated mortality is low, morbidity and economic impact are high. Current licensed vaccines are limited and mediate only partial protection, thus a cost-effective vaccine with improved efficacy is strongly needed.

In this work, recombinant dengue serotype 1 E protein was produced in E. coli, inclusion bodies were isolated and the E protein solubilized in urea and purified using an immobilized metal chelate affinity column. The protein was refolded by dialysis in order to obtain virus-like particles (VLPs).

Particle assembly was confirmed using size-exclusion chromatography, dynamic light scattering (DLS), transmission electron microscopy (TEM), atomic force microscopy and stimulated emission depletion fluorescence (STED) microscopy. Particle diameter was strongly dependent on temperature, pH, buffer salt composition, and addition of L-arginine. Particles were stable in carbonate buffer at pH 9.5 and higher at 4 °C and did not aggregate during short-term temperature increase up to 55 °C.

However, on basis of the above analyses, especially the results of DLS, TEM and STED, it was concluded that the particles obtained did not have an optimal virus-like structure and were therefore designated “virus-sized particles” (VSPs) rather than VLPs.

Immunization of rabbits with the particles did not induce neutralizing antibodies, despite the recognition of the native virus by rabbit antibodies. As the titers against the immunogen were much higher than against the (heat-inactivated) virus, it is assumed that the conformation of the particles at the time of immunization was not optimal. Studies are currently underway to improve the quality of the E protein virus-sized particles towards true virus-like particles in order to optimize its potential as a dengue vaccine candidate.

登革热是世界上传播最广泛的媒介传播疾病之一。尽管与登革热相关的死亡率很低，但发病率和经济影响却很高。当前许可的疫苗是有限的并且仅介导部分保护，因此强烈需要具有改善的功效的成本有效的疫苗。

在这项工作中，在大肠杆菌中生产了重组登革热血清型1 E蛋白，分离了包涵体，将E蛋白溶解在尿素中，并使用固定的金属螯合亲和柱纯化。通过透析使蛋白质重折叠以获得病毒样颗粒（VLP）。

使用尺寸排阻色谱法，动态光散射（DLS），透射电子显微镜（TEM），原子力显微镜和受激发射耗尽荧光（STED）显微镜确认颗粒组装。粒径在很大程度上取决于温度，pH，缓冲盐组成和L-精氨酸的添加。颗粒在pH 9.5的碳酸盐缓冲液中稳定，在4°C时更高，并且在短期温度升高到55°C时不会聚集。

但是，根据上述分析，尤其是DLS，TEM和STED的结果，得出的结论是，所获得的颗粒不具有最佳的病毒样结构，因此被指定为“病毒大小的颗粒”（VSP），而不是VLP。

尽管用兔抗体识别了天然病毒，但是用颗粒免疫兔并没有诱导中和抗体。由于针对免疫原的效价远高于针对（热灭活）病毒的效价，因此可以认为免疫时颗粒的构象不是最佳的。目前正在进行研究以将E蛋白病毒大小的颗粒的质量提高为真正的病毒样颗粒，以优化其作为登革热疫苗候选者的潜力。