Development of a rapid neutralization assay for the detection of neutralizing antibodies against coxsackievirus B1

Yin, Zhichao; Zhu, Rui; Xu, Longfa; Huang, Qiongzi; Zhang, Dongqing; Yang, Hongwei; Zhou, Zhenqi; Zhang, Jun; Cheng, Tong; Xia, Ningshao

doi:10.1016/j.diagmicrobio.2022.115676

Cited by 2 publications

(1 citation statement)

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“…In 2021, a novel quantitative PCR-based assay for detecting NAbs against several RV serotypes was reported by Choi et al However, this method is relatively complex and inefficient, and can only be conducted in 24-well cell culture plates, requiring the extraction of viral nucleotides and RT-PCR after 3 days of incubation (Choi et al, 2021). Recently, an ELISPOT-based neutralization assay (Nt-ELISPOT) has been developed and it is widely used for the detection of NAb levels against many types of viruses, such as SARS-CoV-2 (Park et al, 2021), CVB1 (Wu et al, 2022), CVA10 (Liu et al, 2019), ZIKA (Li et al, 2020), and HSV-1 (Luo et al, 2016). The established Nt-ELISPOT showed characteristics such as high sensitivity, objectivity, efficiency, and is high throughput.…”

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confidence: 99%

Development of a rapid neutralization testing system for Rhinovirus C15 based on the enzyme-linked immunospot assay

et al. 2022

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Human Rhinoviruses (RVs) are dominant pathogens causing a wide range of respiratory tract diseases, posing a huge threat to public health worldwide. Viruses belonging to the RV-C species are more likely to cause severe illnesses and are strongly associated with asthma onset or exacerbations than RV-A or RV-B. Rapid and sensitive detection of neutralizing antibodies (NAbs) against RV-C can promote the development of vaccines and antiviral drugs and help in the diagnosis of viral infection. In this study, a rapid neutralization testing system for RV-C15, based on an enzyme-linked immunospot assay (Nt-ELISPOT) was developed. A monoclonal antibody (MAb), named 9F9, with high binding efficacy for RV-C15 conjugated to horseradish peroxidase (HRP), was used to detect RV-C15-infected cells at a concentration of 2 μg/ml. The optimal infectious dose of RV-C15 was set at 1 × 104 TCID50/well and the cells were fixed with 0.5% formaldehyde diluted in PBS after incubation for 20 h. Compared with the traditional cytopathic effect (CPE)-based neutralization assay (Nt-CPE), Nt-ELISPOT significantly shortened the detection period and showed good consistency with the detection of neutralizing titers of both sera and NAbs. Using Nt-ELISPOT, three anti-RV-C15 NAbs were obtained with IC50 values of 0.16, 0.27, and 11.8 μg/ml, respectively. Moreover, 64 human serum samples collected from a wide range of age groups were tested for NAb against RV-C15 by Nt-ELISPOT. The total seroprevalence was 48.4% (31/64) and the positive rate was lowest in the group under 6 years old. Thus, the Nt-ELISPOT established in this study can be used as a high-throughput and rapid neutralization assay for the screening of NAbs and for seroepidemiological investigation against RV-C15.

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