Development of an indirect enzyme-linked immunosorbent assay (ELISA) assay based on a recombinant truncated VP2 (tVP2) protein for the detection of canine parvovirus antibodies

Shi, Lisong; Wang, Jing; Wang, Peng; Li, Gang; Gong, Miaomiao; Yuan, Weifeng; Zhu, Hongfei

doi:10.5897/ajb12.2446

Cited by 7 publications

(4 citation statements)

References 24 publications

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“…The recombinant full-length and 35 kDa fragment VP2 proteins in the present study were insoluble, as observed in other studies that used E. coli as the VP2 expression host [ 9 , 22 ]. However, this had no effect on the ability of the recombinant VP2 protein to interact with polyclonal antibodies against CPV [ 9 , 22 - 24 ], which was similar to the present study. Insoluble recombinant proteins are generated due to incorrectly folded proteins and stabilized by removing hydrophobic residues [ 25 ].…”

Section: Discussionsupporting

confidence: 89%

“…In this study, the recombinant full-length and the 35 kDa fragment VP2 proteins interacted specifically with rabbit anti-CPV polyclonal antibodies, even though there are 10 antigenic sites in the recombinant full-length VP2 protein and only seven antigenic sites in the recombinant 35 kDa fragment VP2 protein; this may have been due to the high immunoactivity of the VP2 protein of CPV located on loops 1 and 3 [ 12 , 14 ] that contain epitopes 1-7. In addition, the recombinant truncated VP2 protein containing epitope 5 or epitope 6-7 expression in E. coli has been shown to interact with canine serum vaccinated with CPV vaccine [ 24 ]. These recombinant truncated VP2 proteins are good candidates for ELISA or latex agglutination tests for CPV.…”

Section: Discussionmentioning

confidence: 99%

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Expression of recombinant 35 kDa fragment of VP2 protein of canine parvovirus using Escherichia coli expression system

et al. 2021

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Background and Aim: Canine parvovirus (CPV) is one of the most common viral infections in dogs, causing acute hemorrhagic gastroenteritis and high mortality. Vaccination effectively prevents CPV infection. However, the currently available CPV vaccines have concerns such as maternal immunity interference, shedding of virus vaccine, and false-positive result based on polymerase chain reaction after vaccination. A subunit vaccine can overcome these problems. This study aimed to express the recombinant 35 kDa fragment of the VP2 protein (consisting of epitopes 1-7) and the recombinant full-length VP2 protein (consisting of epitopes 1-10) and to study the ability of these two recombinant proteins to react with rabbit anti-CPV polyclonal antibodies. Materials and Methods: The full length and 35 kDa fragment of VP2 gene of CPV were cloned into the pBAD202 Directional TOPOTM expression vector and expressed in E. coli. The recombinant full-length and the recombinant 35 kDa fragment proteins of VP2 were analyzed using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting. Results: The recombinant full-length and the recombinant 35 kDa fragment VP2 genes were successfully cloned and expressed. The optimum concentrations of arabinose and induction time for the recombinant full-length and the recombinant 35 kDa fragment VP2 proteins were 0.2% for 6 h and 0.02% for 6 h, respectively. The recombinant full-length and the recombinant 35 kDa fragment VP2 protein molecular weights were approximately 81 and 51 kDa, respectively. The recombinant full-length and the recombinant 35 kDa fragment VP2 proteins specifically interacted with rabbit anti-CPV polyclonal antibodies. Conclusion: These results suggest that the recombinant 35 kDa fragment and the recombinant full-length VP2 proteins may be useful in developing a CPV diagnostic test or vaccine.

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Section: Discussionsupporting

confidence: 89%

Section: Discussionmentioning

confidence: 99%

Expression of recombinant 35 kDa fragment of VP2 protein of canine parvovirus using Escherichia coli expression system

et al. 2021

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“…The polypeptide consisting of epitopes (EP123) did not give positive result in any dilution. In order for an antigen to be considered immunogenic, it must be positive at a dilution of up to 1/160 [42]. According to these results, only VP1 can be considered as an immunogen (Fig.…”

Section: Immunogenicity Levels Of Vp1 Vp2 P22and Ep123 Recombinant Pr...mentioning

confidence: 99%

Molecular Cloning and Immunogenicity Determination of Norovirus Proteins as Vaccine Candidates

Bingül,

Başbülbül

2024

Preprint

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Human Noroviruses (HuNoVs) are considered the main cause of gastroenteritis in developed and developing countries. Aim of this research was to recombinant production of some structural and functional Norovirus proteins and to determine their immunogenicity in mice. Synthetic VP1, VP2, p22 and a polypeptide (EP123) sequences were amplified with PCR and then amplicons in pET-30a (+) expression vector were transformed into E. coli BL21 cells. Recombinantly produced proteins were purified by Ni-NTA chromotograhy and ammonium sulphate precipitation. Molecular weights of recombinant VP1, VP2, P22 and EP123 were estimated as 63, 34.4, 26 and 27.9 kDa, respectively. Indirect ELISA method was applied to detect IgG levels from serum samples of vaccinated mice. Considering that samples with a p/n ratio of 2 and greater than 2 were positive, VP1 was found to be immunogenic up to a dilution of 1/160 (p/n = 2.09). While VP2 and P22 were found to be immunogenic up to a dilution of 1/80 and 1/20 respectively, EP123 did not give positive result in any dilution. These results suggest that recombinantly produced VP1, has immunogenic potential, whereas VP2, P22 and EP123 polypeptide did not show promising result as a vaccine candidate.

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“…Therefore, the recombinant protein based diagnostic tests can be used as alternative methods for detection of CPV infection. Recombinant VP2 protein-based indirect ELISA assay was found to be economical and more convenient (Lijun et al, 2012). New CPV-2a strain was found to be prevalent CPV strain circulating in India (Mukhopadhyay et al, 2013;Mittal et al, 2014), thus the development of a diagnostic tool based on the prevailing antigenic strain is the need of the hour.…”

Section: Article Infomentioning

confidence: 99%

Cloning and Expression of Recombinant VP2 Capsid Protein Gene of Canine Parvovirus in E. coli System

Sangeetha¹,

Mukhopadhyay²,

Srinivas³

et al. 2018

Int.J.Curr.Microbiol.App.Sci

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Development of an indirect enzyme-linked immunosorbent assay (ELISA) assay based on a recombinant truncated VP2 (tVP2) protein for the detection of canine parvovirus antibodies

Cited by 7 publications

References 24 publications

Expression of recombinant 35 kDa fragment of VP2 protein of canine parvovirus using Escherichia coli expression system

Expression of recombinant 35 kDa fragment of VP2 protein of canine parvovirus using Escherichia coli expression system

Molecular Cloning and Immunogenicity Determination of Norovirus Proteins as Vaccine Candidates

Cloning and Expression of Recombinant VP2 Capsid Protein Gene of Canine Parvovirus in E. coli System

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