PDF(Pubmed)
Abstract：
Virus-specific antibodies are crucial for protective immunity against SARS-CoV-2. Assessing functional antibodies through conventional or pseudotyped virus neutralisation tests (pVNT) requires high biosafety levels. Alternatively, the virus-free surrogate virus neutralisation test (sVNT) quantifies antibodies interfering with spike binding to angiotensin-converting enzyme 2. We evaluated secreted nanoluciferase-tagged spike protein fragments as diagnostic antigens in the sVNT in a vaccination cohort. Initially, spike fragments were tested in a capture enzyme immunoassay (EIA), identifying the receptor binding domain (RBD) as the optimal diagnostic antigen. The sensitivity of the in-house sVNT applying the nanoluciferase-labelled RBD equalled or surpassed that of a commercial sVNT (cPass, GenScript Diagnostics) and an in-house pVNT four weeks after the first vaccination (98% vs. 94% and 72%, respectively), reaching 100% in all assays four weeks after the second and third vaccinations. When testing serum reactivity with Omicron BA.1 spike, the sVNT and pVNT displayed superior discrimination between wild-type- and variant-specific serum reactivity compared to a capture EIA. This was most pronounced after the first and second vaccinations, with the third vaccination resulting in robust, cross-reactive BA.1 construct detection. In conclusion, utilising nanoluciferase-labelled antigens permits the quantification of SARS-CoV-2-specific inhibitory antibodies. Designed as flexible modular systems, the assays can be readily adjusted for monitoring vaccine efficacy.
摘要：
病毒特异性抗体对于抗SARS-CoV-2的保护性免疫至关重要。通过常规或假型病毒中和测试(pVNT)评估功能性抗体需要高生物安全水平。或者，无病毒替代病毒中和试验(sVNT)对干扰刺突与血管紧张素转换酶2结合的抗体进行定量.我们评估了分泌的纳米荧光素酶标记的刺突蛋白片段作为疫苗接种队列中sVNT的诊断抗原。最初，在捕获酶免疫测定（EIA）中测试了刺突片段，鉴定受体结合域（RBD）为最佳诊断抗原。内部sVNT应用纳米荧光素酶标记的RBD的灵敏度等于或超过商业sVNT（cPass，GenScript诊断）和首次接种疫苗四周后的内部pVNT（98%与94%和72%，分别)，在第二次和第三次疫苗接种四周后，所有检测都达到100%。用OmicronBA.1尖峰测试血清反应性时，与捕获EIA相比,sVNT和pVNT在野生型和变体特异性血清反应性之间显示出更好的区分度.这在第一次和第二次接种疫苗后最为明显，第三次疫苗接种导致了强大的，交叉反应BA.1构建体检测。总之,利用纳米荧光素酶标记的抗原可以定量SARS-CoV-2特异性抑制性抗体。设计为灵活的模块化系统，可以容易地调整测定以监测疫苗效力。
