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Development of a recombinant protein-based ELISA for detection of antibodies against bovine herpesvirus 6 (BoHV6)

Piotr Kubiś
Piotr Kubiś
 oraz 
Jacek Kuźmak
Jacek Kuźmak
   | 19 gru 2023
Journal of Veterinary Research's Cover Image
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Data publikacji: 19 gru 2023
Zakres stron: 509 - 515
Otrzymano: 08 paź 2023
Przyjęty: 05 gru 2023
DOI: https://doi.org/10.2478/jvetres-2023-0069
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© 2023 Piotr Kubiś et al., published by Sciendo
This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 License.

Fig. 1.

Electrophoresis in 1% agarose gel of nested PCR amplification products of bovine herpesvirus 6 glycoprotein-coding genes. M – DNA size marker GeneRuler 1 kb Plus DNA ladder (Thermo Scientific, Vilnius, Lithuania); NC – negative control; 1 – 1,389 bp fragment of gB gene; 2 – 1,254 bp fragment of gH gene
Electrophoresis in 1% agarose gel of nested PCR amplification products of bovine herpesvirus 6 glycoprotein-coding genes. M – DNA size marker GeneRuler 1 kb Plus DNA ladder (Thermo Scientific, Vilnius, Lithuania); NC – negative control; 1 – 1,389 bp fragment of gB gene; 2 – 1,254 bp fragment of gH gene

Fig. 2.

Electrophoresis of purified recombinant gH (48 kDa) and gB (53 kDa) proteins of bovine herpesvirus 6 in 10% sodium dodecyl sulphate polyacrylamide gel electrophoresis in denaturing conditions. M – PageRuler Prestained Protein Ladder molecular weight marker (Thermo Scientific, Vilnius, Lithuania)
Electrophoresis of purified recombinant gH (48 kDa) and gB (53 kDa) proteins of bovine herpesvirus 6 in 10% sodium dodecyl sulphate polyacrylamide gel electrophoresis in denaturing conditions. M – PageRuler Prestained Protein Ladder molecular weight marker (Thermo Scientific, Vilnius, Lithuania)

Fig. 3.

Western blot detection of recombinant gH (A) and recombinant gB (B) proteins of bovine herpesvirus 6 with bovine serum samples. Lines 1–4 – field positive sera; line 5 – positive control serum; line 6 – negative control serum; lines 7–8 – field negative sera; M – PageRuler Prestained Protein Ladder molecular weight marker (Thermo Scientific, Vilnius, Lithuania)
Western blot detection of recombinant gH (A) and recombinant gB (B) proteins of bovine herpesvirus 6 with bovine serum samples. Lines 1–4 – field positive sera; line 5 – positive control serum; line 6 – negative control serum; lines 7–8 – field negative sera; M – PageRuler Prestained Protein Ladder molecular weight marker (Thermo Scientific, Vilnius, Lithuania)

Fig. 4.

Distribution of sample-to-positive ratio (S/P) values among bovine sera tested for bovine herpesvirus 6 with recombinant glycoprotein B (rgB) antigen
Distribution of sample-to-positive ratio (S/P) values among bovine sera tested for bovine herpesvirus 6 with recombinant glycoprotein B (rgB) antigen

Fig. 5.

Distribution of sample-to-positive ratio (S/P) values among bovine sera tested for bovine herpesvirus 6 with recombinant glycoprotein H (rgH) antigen
Distribution of sample-to-positive ratio (S/P) values among bovine sera tested for bovine herpesvirus 6 with recombinant glycoprotein H (rgH) antigen

Fig. 6.

Scatter plot analysis of ELISA results showing relationship between bovine herpesvirus 6 recombinant gB and recombinant gH (rgB and rgH) antigens. S/P – sample-to-positive ratio
Scatter plot analysis of ELISA results showing relationship between bovine herpesvirus 6 recombinant gB and recombinant gH (rgB and rgH) antigens. S/P – sample-to-positive ratio

Primers1) used in this study for amplification of bovine herpesvirus 6 DNA fragments

Primer Genome position2) Primer sequence (5’→3’) Use
gBF 35446-35467 AACCTTATCCCGTACATGTTTC PCR
gBR 37722–37691 CAAAGACCAACATGCCGCCAAA PCR
gHF 54713–56173 GAGTCTGGCTTGAATGACGATC PCR
gHR 56021–54767 GGGGTCAGTAATGCAGGGCCTA PCR
pL52gBF3) 35668–35689 GGTTGGGAATTGCAATCTAACATCACGGTGGACCTTA nested PCR, cloning
pL52gBR 37056–37036 GGAGATGGGAAGTCATTAGCTGCTTAAGCGCTTCTCTTC nested PCR, cloning
pL52gHF 54767–54787 GGTTGGGAATTGCAATACAAAGTAGACAAAGAAGCT nested PCR, cloning
pL52gHR4) 56021–55090 GGAGATGGGAAGTCATTATGATTCTTTGTCTACTTTGTA nested PCR, cloning
LICfor55) 296–316 TAATACGACTCACTATAGGG sequencing, colony PCR
LICrev 559–538 GAGCGGATAACAATTTCACAGG sequencing, colony PCR
eISSN:
2450-8608
Język:
Angielski
Częstotliwość wydawania:
4 razy w roku
Dziedziny czasopisma:
Life Sciences, Molecular Biology, Microbiology and Virology, other, Medicine, Veterinary Medicine
Kanał RSS czasopisma