Epitope analysis of glycoprotein I of pseudorabies virus

Jacobs, Liesbeth; Meloen, Rob H.; Gielkens, A. L. J.; Oirschot, J.T. van

doi:10.1099/0022-1317-71-4-881

Cited by 34 publications

(34 citation statements)

References 22 publications

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“…It was unexpected that two pigs infected with mutant M303 would develop an antibody titre against gI, because the gI ELISA specifically detects antibodies directed against antigenic domains C and E, which were destroyed by the deletion of valine(125) and cysteine(126) (Jacobs et al, 1990(Jacobs et al, , 1993. It is possible that steric hindrance caused this blocking effect, particularly because blocking occurs only at a low dilution of the pig serum.…”

Section: Discussionmentioning

confidence: 99%

“…Briefly, we constructed two gI mutants by deleting two amino acids in antigenic domains of PRV gI. M303 (A 125,126) had a deletion of amino acids valine(125) and cysteine(126), which resulted in the loss of the discontinuous antigenic domains C and E. M304 (A59,60) had a deletion of amino acids glycine(59) and aspartic acid(60) which resulted in the loss of the continuous antigenic domain B (Jacobs et al, 1990). In addition, a gI strain (M301) was constructed by the introduction of 10 nucleotides at amino acid position 124, which resulted in a frameshift.…”

Section: Methodsmentioning

confidence: 99%

“…One had a deletion of valine(125) and cysteine(126) (termed M303; A125,126). The other had a deletion of glycine(59) and aspartic acid(60) (termed M304; A59,60) (Jacobs et al, 1990). A gI strain (M301) was constructed by the introduction of a frameshift.…”

Section: Introductionmentioning

confidence: 99%

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Deleting two amino acids in glycoprotein gI of pseudorabies virus decreases virulence and neurotropism for pigs, but does not affect immunogenicity

Jacobs¹,

Mulder²,

Oirschot³

et al. 1993

Journal of General Virology

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The virulence, pathogenicity and immunogenicity of two pseudorabies virus (PRV) variants were investigated in 3-week-old pigs that had been intranasally infected. Variant M303 (A125,126) lacked amino acids valine (125) and cysteine(126) in an immunodominant antigenic region of glycoprotein I (gI) containing two discontinuous antigenic domains, whereas M304 (A59,60) lacked amino acids glycine(59) and aspartic acid(60) in a continuous antigenic domain. M303 (A125,126) was not virulent for pigs, but M304 (A59,60) was as virulent as wild-type PRV: all pigs died within 8 days of infection. Both gI mutant viruses replicated in the oropharyngeal mucosa, although M304 (A59,60) replicated to higher virus titres than M303 (A125,126), and virus was recovered from various tissues. However, in contrast to M304 (A59,60), M303 (A125,126) was not recovered from any central nervous system (CNS) tissues examined. Thus, the tendency of PRV to locate in the CNS was markedly reduced by deleting amino acids valine(125) and cysteine(126) ofgI. Pigs immunized with M303 were completely protected against challenge infection; no clinical signs of disease were detected, no virus was shed, and no secondary antibody response was detected. Thus, deleting amino acids valine(125) and cysteine(126) in gI decreases virulence and neurotropism and does not affect immunogenicity.

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Deleting two amino acids in glycoprotein gI of pseudorabies virus decreases virulence and neurotropism for pigs, but does not affect immunogenicity

Jacobs¹,

Mulder²,

Oirschot³

et al. 1993

Journal of General Virology

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“…Although the N-terminal part of gE comprises a cluster of conformation-independent epitopes and is considered to be one of the most important immunodominant regions of the glycoprotein (Fuchs et al, 1990;Jacobs et al, 1990;Morenkov et al, 1997=), the cloned recombinant protein exhibited a relatively weak reactivity with sera from infected animals as compared to the affinity-purified gE. This is most probably connected with the lack of immunodominant conformationdependent epitopes on the recombinant gE protein that induce a strong antibody response during infection of swine.…”

Section: Discussionmentioning

confidence: 99%

Indirect Elisas Based on Recombinant and Affinity-Purified Glycoprotein E of Aujeszky's Disease Virus to Differentiate Between Vaccinated and Infected Animals

Morenkov

Fodor

1999

Acta Veterinaria Hungarica

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Two indirect ELISAs for the detection of antibodies against glycoprotein E (gE) of Aujeszkys disease virus (ADV) in sera have been developed. The rec-gE-ELISA is based on the E. coli-expressed recombinant protein containing the Nterminal sequences of gE (aa 1-125) fused with the glutathione S-transferase from Schistosoma japonicum. The affi-gE-ELISA is based on native gE, which was purified from virions by affinity chromatography. The tests were optimised and compared with each other, as well as with the recently developed blocking gE-ELISA (Morenkov et al., 1997>), with respect to specificity and sensitivity. The recgE-ELISA was less sensitive in detecting ADV-infected animals than the affi-gE-ELISA (sensitivity 80% and 97%, respectively), which is probably due to the lack of conformation-dependent immunodominant epitopes on the recombinant protein expressed in E. coli. The specificity of the rec-gE-ELISA and affi-gE-ELISA was rather moderate (90% and 94%, respectively) because it was necessary to set such cut-off values in the tests that provided a maximum level of sensitivity, which obviously increased the incidence of false positive reactions. Though the indirect ELISAs detect antibodies against many epitopes of gE, the blocking gE-ELISA, which detects antibodies against only one immunodominant epitope of gE, showed a better test performance (specificity 99% and sensitivity 98%). This is most probably due to rather high dilutions of the sera used in the indirect gE-ELISAs (1:30) as compared to the serum dilution in the blocking gE-ELISA (1:2). We conclude that the indirect gE-ELISAs are sufficiently specific and sensitive to distinguish ADVinfected swine from those vaccinated with gE-negative vaccine and can be useful, in particularly affi-gE-ELISA, as additional tests for the detection of antibodies to gE.

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“…(7) It is hard to obtain complete gE by expressing full length gE gene in heterologous cells. It is therefore preferable to prepare high affinitive monoclonal antibodies against PRV by using inactivated whole virus as immunogen.…”

Section: Introductionmentioning

confidence: 99%

Immune-Tolerizing Procedure for Preparation of Monoclonal Antibodies Against Glycoprotein E of Pseudorabies Virus

Xiang

Dan

et al. 2013

Monoclonal Antibodies in Immunodiagnosis and Immunotherapy

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The glycoprotein E (gE) of pseudorabies virus (PRV) is known to be an important marker protein in the control and eradication of Aujeszky's disease. In this study, BALB/c mice were immunized with gE-deleted PRV as tolerogen and with wild-type PRV as immunogen. The spleen cells from the immunized mice were then fused with the myeloma cell line Sp2/0. Two hybridoma cell lines that could stably secrete the monoclonal antibody (MAb) against gE were achieved by using indirect ELISA screening and subcloning three times; they were named 1D2 and 2B2. Indirect immunofluorescence assay (IFA) revealed that the MAbs were specifically against gE of PRV. MAbs 1D2 and 2B2 were subgroup IgG1. The MAbs obtained in this study provide useful tools for the development of differential diagnostic methods for PRV.

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Epitope analysis of glycoprotein I of pseudorabies virus

Cited by 34 publications

References 22 publications

Deleting two amino acids in glycoprotein gI of pseudorabies virus decreases virulence and neurotropism for pigs, but does not affect immunogenicity

Deleting two amino acids in glycoprotein gI of pseudorabies virus decreases virulence and neurotropism for pigs, but does not affect immunogenicity

Indirect Elisas Based on Recombinant and Affinity-Purified Glycoprotein E of Aujeszky's Disease Virus to Differentiate Between Vaccinated and Infected Animals

Immune-Tolerizing Procedure for Preparation of Monoclonal Antibodies Against Glycoprotein E of Pseudorabies Virus

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