PDF(Pubmed)
Abstract：
Coronavirus disease of 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is an ongoing outbreak, disrupting human life worldwide. Vaccine development was prioritized to obtain a biological substance for combating the viral pathogen and lessening disease severity. In vaccine production, biological origin and relevant materials must be carefully examined for potential contaminants in conformity with good manufacturing practice. Due to fast mutation, several SARS-CoV-2 variants and sublineages have been identified. Currently, most of COVID-19 vaccines are developed based on the protein sequence of the Wuhan wild type strain. New vaccines specific for emerging SARS-CoV-2 strains are continuously needed to tackle the incessant evolution of the virus. Therefore, in vaccine development and production, a reliable method to identify the nature of subunit vaccines is required to avoid cross-contamination. In this study, liquid chromatography-mass spectrometry using quadrupole-time of flight along with tryptic digestion was developed for distinguishing protein materials derived from different SARS-CoV-2 strains. After analyzing the recombinantly produced receptor-binding domain (RBD) of the SARS-CoV-2 spike protein, nine characteristic peptides were identified with acceptable limits of detection. They can be used together to distinguish 14 SARS-CoV-2 strains, except Kappa and Epsilon. Plant-produced RBD-Fc protein derived from Omicron strains can be easily distinguished from the others with 4-5 unique peptides. Eventually, a peptide key was developed based on the nine peptides, offering a prompt and precise flowchart to facilitate SARS-CoV-2 strain identification in COVID-19 vaccine manufacturing.
摘要：
2019年冠状病毒病（COVID-19），由严重急性呼吸综合征冠状病毒2（SARS-CoV-2）引起，是一个持续的爆发，扰乱全世界的人类生活。疫苗开发被优先考虑以获得用于对抗病毒病原体和减轻疾病严重程度的生物物质。在疫苗生产中,生物来源和相关材料必须仔细检查潜在的污染物符合良好的生产规范。由于快速突变,已经鉴定了几种SARS-CoV-2变体和亚谱系。目前,大多数COVID-19疫苗是基于武汉野生型菌株的蛋白质序列开发的。不断需要针对新兴SARS-CoV-2毒株的新疫苗来应对病毒的不断进化。因此,在疫苗开发和生产中，为了避免交叉污染，需要一种可靠的方法来鉴定亚单位疫苗的性质。在这项研究中,开发了使用四极杆飞行时间和胰蛋白酶消化的液相色谱-质谱法，用于区分来自不同SARS-CoV-2菌株的蛋白质材料。在分析了SARS-CoV-2刺突蛋白的重组产生的受体结合域（RBD）后，鉴定出9种特征肽，检测限可接受。它们可以一起用于区分14种SARS-CoV-2菌株，除了Kappa和Epsilon.源自0微米菌株的植物产生的RBD-Fc蛋白可以用4-5种独特的肽容易地与其他蛋白区分开。最终,基于九种肽开发了一个肽键，提供及时准确的流程图，以促进SARS-CoV-2株在COVID-19疫苗生产中的鉴定。
