A monoclonal antibody specific to the non-epitope region of hepatitis B virus preS1 contributes to more effective HBV detection

Grouper iridovirus (GIV) is one of the most important viral pathogens in grouper, particularly at the fry and fingerling stages. The study of GIV pathogenicity has been hampered by the lack of proper immunological reagents to study the expression and function of viral proteins in the infected cells. In this study, two mouse monoclonal antibodies (mAbs) against GIV 55L and 97L proteins were produced. Enzyme-linked immunosorbent assay (ELISA) and Western blotting were used to screen these hybridomas, resulting in the identification of two high-affinity mAbs named GIV55L-mAb-2 and GIV97L-mAb-3, respectively. Both mAbs belong to the IgG1 isotype and were effective in detecting their respective target viral protein. Reverse-transcription polymerase chain reaction (RT-PCR) and Western blot analyses of GIV-infected GK cells revealed that GIV 97L is an immediate early gene, whereas GIV 55L a late one. The localization of 55L and 97L in GIV-infected cells was further characterized by immunofluorescence microscopy with the mAbs. The 55L protein mainly aggregated in the cytoplasm while 97L distributed in both the nucleus and cytoplasm of the infected cells. These studies demonstrate the validity of the two mAbs as immunodiagnostic and research reagents.

“…; Zhang et al . ,b). In recent years, mAb technology has had an important impact on aquaculture disease diagnosis (Lai et al .…”

Section: Discussionmentioning

confidence: 99%

Development and characterization of two monoclonal antibodies against grouper iridovirus 55L and 97L proteins

Cheng

et al. 2014

“…; Zhang et al . ). In recent years, mAb technology has had an important impact on aquaculture disease diagnostics (Lai et al .…”

Section: Discussionmentioning

confidence: 97%

“…The evolutionary relationship of GIV-64L with other homologues was consistent with the position of GIV in iridovirus evolution. Monoclonal antibodies are a good tool for detecting and characterizing viral infection (Patil et al 2013;Zhang et al 2013). In recent years, mAb technology has had an important impact on aquaculture disease diagnostics (Lai et al 2001(Lai et al , 2002Shi et al 2003;Cote et al 2009;Hou et al 2011;Aamelfot et al 2013;Patil et al 2013;Siriwattanarat et al 2013).…”

Section: Discussionmentioning

confidence: 99%

Production and characterization of a monoclonal antibody against a late gene encoded by grouper iridovirus 64L

et al. 2015

Viral envelope proteins play important roles in viral infection and assembly. The grouper iridovirus ORF 64L (GIV-64L) was predicted to encode an envelope protein and was conserved in all sequenced Ranaviruses. In this study, the complete nucleotide sequence of the GIV-64L gene (1215 bp) was cloned into the isopropyl β-D-1-thiogalactopyranoside (IPTG) induction prokaryotic expression vector pET23a. The approximately 50.2 kDa recombinant GIV-64L-His protein was induced, purified and used as an immunogen to immunize BALB/c mice. Three monoclonal antibodies (mAbs), all IgG1 class antibodies against GIV-64L protein, were produced by enzyme-linked immunosorbent assay. Reverse transcription polymerase chain reaction analyses revealed GIV-64L to be a late gene when expressed in grouper kidney cells during GIV infection with cycloheximide (an inhibitor of protein synthesis) or cytosine arabinoside (an inhibitor of DNA synthesis) present. Finally, one of the established mAbs, GIV-64L-mAb-17, was used in Western blotting and an immunofluorescence assay, which showed that GIV-64L protein was expressed at 24 h post-infection and localized only in the cytoplasm in GIV-infected cells, packed into a whole virus particle. The presently characterized GIV-64L mAbs should have widespread applications in GIV immunodiagnostics and other research, and these results should offer important insights into the pathogenesis of GIV.

“…; Zhang et al . ). Monoclonal antibody technology has made important contribution to aquaculture disease diagnosis in recent years (Lai et al .…”

Section: Discussionmentioning

confidence: 97%

“…Monoclonal antibodies are a good tool for the detection and characterization of viruses (Patil et al 2013;Zhang et al 2013). Monoclonal antibody technology has made important contribution to aquaculture disease diagnosis in recent years (Lai et al 2001(Lai et al , 2002Shi et al 2003;Cote et al 2009;Hou et al 2011;Aamelfot et al 2013;Patil et al 2013;Siriwattanarat et al 2013).…”

Section: Discussionmentioning

confidence: 99%

Identification and characterization of a late gene encoded by grouper iridovirus 2L (GIV‐2L)

Lin

Cheng

et al. 2014

Grouper iridovirus (GIV) belongs to the Ranavirus genus and is one of the most important viral pathogens in grouper, particularly at the fry and fingerling stages. In this study, we identified and characterized the GIV-2L gene, which encodes a protein of unknown function. GIV-2L is 1242 bp in length, with a predicted protein mass of 46.2 kDa. It displayed significant identity only with members of the Ranavirus and Iridovirus genera. We produced mouse monoclonal antibodies against the GIV-2L protein by immunizing mice with GIV-2L-His-tag recombinant protein. By inhibiting de novo protein and DNA synthesis in GIV-infected cells, we showed that GIV-2L was a late gene during the viral replication. Finally, immunofluorescence microscopy revealed that GIV-2L protein accumulated in both the nucleus and cytoplasm of infected cells. These results offer important insights into the pathogenesis of GIV.