The receptor-binding domain (RBD) of the SARS-CoV-2 spike protein is a commonly used antigen for serology assays critical to determining the extent of SARS-CoV-2 exposure in the population. Different versions of the RBD protein have been developed and utilized in assays, with higher sensitivity attributed to particular forms of the protein. To improve the yield of these high-sensitivity forms of RBD and support the increased demand for this antigen in serology assays, we investigated several protein expression variables including DNA elements such as promoters and signal peptides, cell culture expression parameters, and purification processes. Through this investigation, we developed a simplified and robust purification strategy that consistently resulted in high levels of the high-sensitivity form of RBD and demonstrated that a carboxyterminal tag is responsible for the increased sensitivity in the ELISA. These improved reagents and processes produce high-quality proteins which are functional in serology assays and can be used to investigate seropositivity to SARS-CoV-2 infection.

SARS-CoV-2 刺突蛋白的受体结合域 (RBD) 是血清学检测中常用的抗原，对于确定人群中 SARS-CoV-2 暴露的程度至关重要。不同版本的 RBD 蛋白已被开发并用于检测，由于该蛋白的特定形式具有更高的灵敏度。为了提高这些高灵敏度 RBD 的产量并支持血清学检测中对该抗原不断增长的需求，我们研究了几种蛋白质表达变量，包括启动子和信号肽等 DNA 元件、细胞培养表达参数和纯化过程。通过这项研究，我们开发了一种简化而强大的纯化策略，该策略始终能够产生高水平的高灵敏度形式的 RBD，并证明羧基末端标签是提高 ELISA 灵敏度的原因。这些改进的试剂和工艺可产生高质量的蛋白质，这些蛋白质可在血清学检测中发挥作用，并可用于研究 SARS-CoV-2 感染的血清阳性反应。