Diagnosing Zika virus (ZIKV) infections has been challenging due to the cross-reactivity of induced antibodies with other flavivirus. The concomitant occurrence of ZIKV and Dengue virus (DENV) in endemic regions requires diagnostic tools with the ability to distinguish these two viral infections. Recent studies demonstrated that immunoassays using the C-terminal fragment of ZIKV NS1 antigen (ΔNS1) can be used to discriminate ZIKV from DENV infections. In order to be used in serological tests, the expression/solubility of ΔNS1 and growth of recombinant E. coli strain were optimized by Response Surface Methodology. Temperature, time and IPTG concentration were evaluated. According to the model, the best condition determined in small scale cultures was 21 °C for 20 h with 0.7 mM of IPTG, which predicted 7.5 g/L of biomass and 962 mg/L of ΔNS1. These conditions were validated and used in a 6-L batch in the bioreactor, which produced 6.4 g/L of biomass and 500 mg/L of ΔNS1 in 12 h of induction. The serological ELISA test performed with purified ΔNS1 showed low cross-reactivity with antibodies from DENV-infected human subjects. Denaturation of ΔNS1 decreased the detection of anti-ZIKV antibodies, thus indicating the contribution of conformational epitopes and confirming the importance of properly folded ΔNS1 for the specificity of the serological analyses. Obtaining high yields of soluble ΔNS1 supports the viability of an effective serologic diagnostic test capable of differentiating ZIKV from other flavivirus infections.

中文翻译：

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大肠杆菌中寨卡病毒ΔNS1的优化和规模化生产：响应面方法学的应用。

由于诱导的抗体与其他黄病毒的交叉反应性，诊断寨卡病毒（ZIKV）感染一直具有挑战性。ZIKV和登革热病毒（DENV）在地方性地区的同时发生需要诊断工具能够区分这两种病毒感染。最近的研究表明，使用ZIKV NS1抗原（ΔNS1）的C末端片段进行的免疫分析可用于将ZIKV与DENV感染区分开。为了用于血清学测试，通过响应表面方法优化了ΔNS1的表达/溶解度和重组大肠杆菌菌株的生长。评价温度，时间和IPTG浓度。根据该模型，在小规模培养中确定的最佳条件是21°C 20 h，使用0.7 mM IPTG，这将预测7.5 g / L的生物量和962 mg / L的ΔNS1。这些条件已得到验证，并在生物反应器中以6升批次使用，在诱导12小时内产生了6.4 g / L的生物量和500 mg / L的ΔNS1。用纯化的ΔNS1进行的血清ELISA试验显示与来自DENV感染的人类受试者的抗体的交叉反应性低。ΔNS1的变性降低了抗ZIKV抗体的检测，因此表明构象表位的贡献，并确认了正确折叠的ΔNS1对于血清学分析特异性的重要性。获得高产量的可溶性ΔNS1支持能够将ZIKV与其他黄病毒感染区分开的有效血清学诊断测试的可行性。用纯化的ΔNS1进行的血清ELISA试验显示与来自DENV感染的人类受试者的抗体的交叉反应性低。ΔNS1的变性降低了抗ZIKV抗体的检测，因此表明构象表位的贡献，并确认了正确折叠的ΔNS1对于血清学分析特异性的重要性。获得高产量的可溶性ΔNS1支持能够将ZIKV与其他黄病毒感染区分开的有效血清学诊断测试的可行性。用纯化的ΔNS1进行的血清ELISA试验显示与来自DENV感染的人类受试者的抗体的交叉反应性低。ΔNS1的变性降低了抗ZIKV抗体的检测，因此表明构象表位的贡献，并确认了正确折叠的ΔNS1对于血清学分析特异性的重要性。获得高产量的可溶性ΔNS1支持能够将ZIKV与其他黄病毒感染区分开的有效血清学诊断测试的可行性。因此表明构象表位的贡献，并证实适当折叠的ΔNS1对于血清学分析的特异性的重要性。获得高产率的可溶性ΔNS1支持能够将ZIKV与其他黄病毒感染区分开的有效血清学诊断测试的可行性。因此表明构象表位的贡献，并证实正确折叠的ΔNS1对于血清学分析特异性的重要性。获得高产量的可溶性ΔNS1支持能够将ZIKV与其他黄病毒感染区分开的有效血清学诊断测试的可行性。