Abstract
Human parechovirus type 3 (HPeV3) is an etiologic agent of respiratory diseases, meningitis, and sepsis-like illness in both infants and adults. Monoclonal antibodies (mAbs) can be a promising diagnostic tool for antigenic diseases such as virus infection, as they offer a high specificity toward a specific viral antigen. However, to date, there is no specific mAb available for the diagnosis of HPeV3 infection. In this study, we developed and characterized mAbs specific for HPeV3 capsid protein VP0. We used cell-free, wheat germ-synthesized viral VP0 protein for immunizing BALB/c mice to generate hybridomas. From the resultant hybridoma clones, we selected nine clones producing mAbs reactive to the HPeV3-VP0 antigen, based on enzyme-linked immunosorbent assay (ELISA). Epitope mapping showed that these mAbs recognized three distinct domains in HPeV3 VP0. Six mAbs recognized HPeV3 specifically and the other three mAbs showed cross-reactivity with other HPeVs. Using the HPeV3-specific mAbs, we then developed an ELISA for viral antigen detection that could be reliably used for laboratory diagnosis of HPeV3. This ELISA system exhibited no cross-reactivity with other related viruses. Our newly developed mAbs would, thus, provide a useful set of tools for future research and ensure HPeV3-specific diagnosis.
Highlights
Human parechoviruses (HPeVs) belong to the Parechovirus genus of the Picornaviridae family [1,2]
Monoclonal antibodies in the hybridoma culture supernatant were tested using enzyme-linked immunosorbent assay (ELISA) with His-tagged recombinant human parechovirus 3 (HPeV3)-VP0 protein
HPeV3-VP0 protein was produced with high aqueous solubility (Figure 1a)
Summary
Human parechoviruses (HPeVs) belong to the Parechovirus genus of the Picornaviridae family [1,2]. The appropriate diagnosis tool for HPeV3 detection might be able to rule out infectious etiology and avoid unnecessary antibiotics use, which is a given because HPeV3 leads to septic shock-like symptoms. Chen et al generated polyclonal antibodies for HPeV3 VP0, and proposed an immunofluorescence-based diagnostic assay [27] This method requires virus isolation by cell culture and takes several weeks for the identification of viral genotype/serotype. Abed et al developed a serological enzyme-linked immunosorbent assay (ELISA), using a synthetic peptide from the VP0 protein of HPeVs [28] It can provide a definitive diagnosis, serological test requires paired serum samples from acute and recovery phases, which makes it difficult to diagnose immediately as a point of care testing (POCT). We obtained nine mAb clones for characterization, and thereafter, generated an ELISA system that is able to detect the HPeV3 VP0 antigens
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