用于检测印度人群中针对严重急性呼吸综合征冠状病毒2（SARS-CoV-2）刺突蛋白、包膜蛋白和膜蛋白抗原的IgG抗体的病毒样颗粒酶联免疫吸附测定法

VLP-ELISA for the Detection of IgG Antibodies against Spike, Envelope, and Membrane Antigens of SARS-CoV-2 in Indian Population.

作者信息

Kumar Dilip, Roy Sourav Singha, Rastogi Ruchir, Arora Kajal, Undale Avinash, Gupta Reeshu, Arora Nupur Mehrotra, Kundu Prabuddha K

机构信息

Research and Developmental Laboratory, Premas Biotech Private Limited, Sector 4, IMT Manesar, Gurgaon 122050, India.

Centre of Research for Development, Parul University, Vadodara 391760, India.

出版信息

Vaccines (Basel). 2023 Mar 27;11(4):743. doi: 10.3390/vaccines11040743.

DOI:10.3390/vaccines11040743

PMID:37112655

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC10145915/

Abstract

BACKGROUND

Serological methods to conduct epidemiological survey are often directed only against the spike protein. To overcome this limitation, we have designed PRAK-03202, a virus-like particle (VLP), by inserting three antigens (Spike, envelope and membrane) of SARS-CoV-2 into a highly characterized -based D-Crypt™ platform.

METHODS

Dot blot analysis was performed to confirm the presence of S, E, and M proteins in PRAK-03202. The number of particles in PRAK-03202 was measured using nanoparticle tracking analysis (NTA). The sensitivity of VLP-ELISA was evaluated in 100 COVID positive. PRAK-03202 was produced at a 5 L scale using fed-batch fermentation.

RESULTS

Dot blot confirmed the presence of S, E, and M proteins in PRAK-03202. The number of particles in PRAK-03202 was 1.21 × 10 mL. In samples collected >14 days after symptom onset, the sensitivity, specificity, and accuracy of VLP-ELISA were 96%. We did not observe any significant differences in sensitivity, specificity, and accuracy when post-COVID-19 samples were used as negative controls compared to pre-COVID-samples. At a scale of 5 L, the total yield of PRAK-03202 was 100-120 mg/L.

CONCLUSION

In conclusion, we have successfully developed an in-house VLP-ELISA to detect IgG antibodies against three antigens of SARS-CoV-2 as a simple and affordable alternative test.

摘要

背景

进行流行病学调查的血清学方法通常仅针对刺突蛋白。为克服这一局限性，我们通过将严重急性呼吸综合征冠状病毒2（SARS-CoV-2）的三种抗原（刺突蛋白、包膜蛋白和膜蛋白）插入到一个高度特征化的基于D-Crypt™的平台中，设计了一种病毒样颗粒（VLP）PRAK-03202。

方法

进行斑点印迹分析以确认PRAK-03202中S、E和M蛋白的存在。使用纳米颗粒跟踪分析（NTA）测量PRAK-03202中的颗粒数量。在100名新冠病毒阳性患者中评估VLP-ELISA的灵敏度。使用补料分批发酵在5升规模下生产PRAK-03202。

结果

斑点印迹证实PRAK-03202中存在S、E和M蛋白。PRAK-03202中的颗粒数量为1.21×10个/毫升。在症状出现14天以上采集的样本中，VLP-ELISA的灵敏度、特异性和准确性为96%。与新冠病毒感染前样本相比，将新冠病毒感染后样本用作阴性对照时，我们未观察到灵敏度、特异性和准确性有任何显著差异。在5升规模下，PRAK-03202的总产量为100 - 120毫克/升。