Background: Lasting immunity to SARS-CoV-2 following infection is questioned because serum antibodies decline in convalescence. However, functional immunity is mediated by long-lived memory T and B (Bmem) cells. Objective: To determine the longevity and immunophenotype of SARS-CoV-2-specific Bmem cells in COVID-19 patients. Methods: Recombinant spike receptor binding domain (RBD) and nucleocapsid protein (NCP) were produced for ELISA-based serology, and biotinylated for fluorescent tetramer generation to identify SARS-CoV-2-specific Bmem cells by flow cytometry with a panel of 13 mAbs. 36 blood samples were obtained from 25 COVID-19 patients (11 paired) between 4-242 days post-symptom onset for detection of neutralizing antibodies, IgG serology and flow cytometry. Results: The recombinant RBD and NCP were specifically recognized by serum IgG in all patients and reactivity declined >20 days post-symptom onset. All patients had detectable RBD- and NCP-specific Bmem cells at 8.23-267.6 cells/ml of blood (0.004-0.13% of B cells) regardless of sampling time. RBD- and NCP-specific Bmem cells predominantly expressed IgM or IgG1, with the latter formed slightly later than the former. RBD-specific IgG+ Bmem were predominantly CD27+, and numbers significantly correlated with circulating follicular helper T cell numbers. Conclusion: RBD- and NCP-specific Bmem cells persisted for 8 months, indicating that the decline in serum antibodies after 1 month does not indicate waning of immunity but a contraction of the immune response. Flowcytometric detection of SARS-CoV-2-specific Bmem cells enables detection of long-term functional immunity following infection or vaccination for COVID-19.

中文翻译：

---

快速和持久的B细胞记忆可产生COVID-19疾病和康复中的SARS-CoV-2尖峰和核衣壳蛋白

背景：感染后对SARS-CoV-2的持久免疫受到质疑，因为血清抗体在恢复过程中下降。但是，功能性免疫是由长寿的记忆T和B（Bmem）细胞介导的。目的：确定SARS-CoV-2特异性Bmem细胞在COVID-19患者中的寿命和免疫表型。方法：生产重组刺突受体结合结构域（RBD）和核衣壳蛋白（NCP），用于基于ELISA的血清学检测，并进行生物素化，用于荧光四聚体生成，通过流式细胞仪与一组13个mAb一起鉴定SARS-CoV-2特异性Bmem细胞。在症状发作后的4-242天之间，从25名COVID-19患者（11对）中获得了36份血样，用于检测中和抗体，IgG血清学和流式细胞仪。结果：重组RBD和NCP在所有患者中均被血清IgG特异性识别，并且在症状发作后> 20天反应性下降。所有患者均具有可检测的RBD和NCP特异的Bmem细胞，无论采样时间如何，其血液浓度为8.23-267.6细胞/ ml（B细胞的0.004-0.13％）。RBD和NCP特异性Bmem细胞主要表达IgM或IgG1，后者的形成要比前者稍晚。RBD特异性IgG + Bmem主要为CD27 +，其数量与循环性滤泡辅助性T细胞数量显着相关。结论：RBD和NCP特异性Bmem细胞持续存在8个月，这表明1个月后血清抗体的下降并不表示免疫力下降，而是免疫反应的收缩。