A cell line expressing the CD2v protein of ASFV was generated. The efficient expression of CD2v protein was determined by immunofluorescence and Western blotting. The CD2v protein was Ni-affinity purified from the supernatant of cell cultures. The CD2v-expressing cells showed properties of hemadsorption, and the secreted CD2v protein exhibited hemagglutinating activity. The antigenicity and immunoprotection ability of CD2v were evaluated by immunizing pigs alone, combined with a cell-line-expressed p30 protein or triple combined with p30 and K205R protein. Immunized pigs were challenged with the highly virulent ASFV strain HLJ/18. Virus challenge results showed that CD2v immunization alone could provide partial protection at the early infection stage. Protein p30 did not show synergistic protection effects in immunization combined with CD2v. Interestingly, immunization with the triple combination of CD2V, p30 and K205R reversed the protection effect. The viremia onset time was delayed, and one pig out of three recovered after the challenge. The pig recovered from ASFV clinical symptoms, the rectal temperature returned to normal levels and the viremia was cleared. The mechanism of this protection effect warrants further investigation.

African swine fever caused by African swine fever virus (ASFV) is an acute, highly contagious swine disease with high mortality. To facilitate effective vaccine development and find more serodiagnostic targets, fully exploring the ASFV antigenic proteins is urgently needed. In this study, the MGF_110-13L was identified as an immunodominant antigen among the seven transmembrane proteins. The main outer-membrane domain of MGF_110-13L was expressed and purified. Two monoclonal antibodies (mAbs; 8C3, and 10E4) against MGF_110-13L were generated. The epitopes of two mAbs were preliminary mapped with the peptide fusion proteins after probing with mAbs by enzyme-linked immunosorbent assay (ELISA) and Western blot. And the two target epitopes were fine-mapped using further truncated peptide fusion protein strategy. Finally, the core sequences of mAbs 8C3 and 10E4 were identified as ⁴⁸WDCQDGICKNKITESRFIDS⁶⁷, and ¹²²GDHQQLSIKQ¹³¹, respectively. The peptides of epitopes were synthesized and probed with ASFV antibody positive pig sera by a dot blot assay, and the results showed that epitope 10E4 was an antigenic epitope. The epitope 10E4 peptide was further evaluated as a potential antigen for detecting ASFV antibodies. To our knowledge, this is the first report of antigenic epitope information on the antigenic MGF_110-13L protein of ASFV.

Japanese encephalitis virus (JEV) is a zoonotic pathogen transmitted primarily by Culex mosquitoes and causes severe neurological diseases in humans and animals. The main endemic areas are the Western Pacific and Southeast Asia, and its geographical distribution has expanded in recent years. The development of a diagnosis for orthoflavivirus infections is hampered by two main problems: the short duration of viremia, resulting in a narrow detection window, and severe cross-reactivity. NS1, a secreted nonstructural protein of orthoflavivirus, holds promise as a new target for overcoming these limitations. In this study, we established a highly specific and sensitive capture ELISA for the JEV NS1 protein. First, the JEV NS1 protein was successfully expressed in mammalian cells and purified using affinity chromatography. Seven mAbs recognizing JEV NS1 were generated, and the mAb 20B6 exhibited the strongest binding affinity. Based on 20B6, a capture ELISA was developed with an optimal coating concentration of 3 μg/mL and an optimal detection antibody working concentration of 0.432 μg/mL. No cross-reactivity was observed with other orthoflaviviruses (including WNV, KUNV, USUV, MVEV, SLEV, and ZIKV) or common porcine viruses. The method could effectively detect NS1 protein in cell culture medium, cell lysates, mouse tissues, and porcine serum samples from JEV-infected subjects. This study provides a solid foundation that may be further developed into an efficient and specific tool for epidemiological surveillance of Japanese encephalitis.