scholarly journals Use of Heavy Water (D2O) in Developing Thermostable Recombinant p26 Protein Based Enzyme-Linked Immunosorbent Assay for Serodiagnosis of Equine Infectious Anemia Virus Infection

2014 ◽  
Vol 2014 ◽  
pp. 1-6
Author(s):  
Harisankar Singha ◽  
Sachin K. Goyal ◽  
Praveen Malik ◽  
Raj K. Singh
Keyword(s):  
Immunosorbent Assay ◽  
Heavy Water ◽  
Equine Infectious Anemia Virus ◽  
Thermal Protection ◽  
Deuterium Oxide ◽  
Influenza Virus Vaccine ◽  
Enzyme Linked Immunosorbent Assay ◽  
Equine Infectious Anemia ◽  
Elisa Plate ◽  
Anemia Virus

Thermostabilizing effect of heavy water (D2O) or deuterium oxide has been demonstrated previously on several enzymes and vaccines like oral poliovirus vaccine and influenza virus vaccine. In view of the above observations, effect of heavy water onin situthermostabilization of recombinant p26 protein on enzyme-linked immunosorbent assay (ELISA) for serodiagnosis of equine infectious anemia virus (EIAV) infection was investigated in the present study. The carbonate-bicarbonate coating buffer was prepared in 60% and 80% D2O for coating the p26 protein in 96-well ELISA plate and thermal stability was examined at 4°C, 37°C, 42°C, and 45°C over a storage time from 2 weeks to 10 months. A set of positive serum (n=12) consisting of strong, medium, and weak titer strength (4 samples in each category) and negative serum (n=30) were assessed in ELISA during the study period. At each time point, ELISA results were compared with fresh plate to assess thermal protective effect of D2O. Gradual increase in the stabilizing effect of 80% D2O at elevated temperature (37°C < 42°C < 45°C) was observed. The 80% D2O provides the thermal protection to rp26 protein in ELISA plate up to 2 months of incubation at 45°C. The findings of the present study have the future implication of adopting cost effective strategies for generating more heat tolerable ELISA reagents with extended shelf life.

scholarly journals Identification and genetic characterization of equine infectious anemia virus in Western Balkans

2021 ◽  
Vol 17 (1) ◽  
Author(s):  
Diana Lupulovic ◽  
Sara Savić ◽  
Delphine Gaudaire ◽  
Nicolas Berthet ◽  
Živoslav Grgić ◽  
...  
Keyword(s):  
Equine Infectious Anemia Virus ◽  
Clinical Signs ◽  
Full Genome Sequence ◽  
Enzyme Linked Immunosorbent Assay ◽  
Full Genome ◽  
Western Balkans ◽  
Full Genome Sequencing ◽  
Horse Population ◽  
Equine Infectious Anemia ◽  
Anemia Virus

Abstract Background Equine infectious anemia (EIA) is a viral disease, caused by the Equine Infectious Anemia virus (EIAV) belonging to the Retroviridae family, genus Lentivirus. Horses (or equids) infected with EIAV are lifelong carriers and they remain contagious for other horses even in the absence of clinical signs. So far, EIAV infection has been reported among horses in North and South America, France, Germany, Italy, Hungary and Romania, with no publication regarding the presence of EIAV in horses in Serbia. To determine the circulation of EIAV among, approximately, the 5000 horses of the Vojvodina region, northern part of Serbia, 316 serum undergone serological testing for EIA. Then, identification and full genome sequencing using next generation sequencing was performed from one EIA positive horse. Results the 316 sera were tested with 3 different commercial agar gel immunodiffusion (AGID) tests and two different commercial enzyme-linked immunosorbent assay (ELISA). With the three AGID kits, 311 (98.4%) among the 316 tested sera were negative and only five (1.6%) sera were positive for EIA. Some discrepancies were seen for the two ELISA kits tested since one exhibited the same results as AGID test and the second gave 295 sera with negative results, five with a positive result and 16 with doubtful outcome. Phylogenetic analysis performed using the full genome sequence showed that EIAV characterized from a horse in Serbia is different from those identify so fare around the world and form a distinct and separate group together with another EIAV strain. Conclusions This study demonstrate for the first time that EIAV is circulating at a low level in the horse population from the Northern part of Serbia. Interestingly, phylogenetic data indicates that this EIAV from the western Balkan region of Europe belongs to a new cluster.

scholarly journals Serological Method Using Recombinant S2 Protein To Differentiate Equine Infectious Anemia Virus (EIAV)-Infected and EIAV-Vaccinated Horses

2004 ◽  
Vol 11 (6) ◽  
pp. 1120-1129 ◽  
Author(s):  
Sha Jin ◽  
Charles J. Issel ◽  
Ronald C. Montelaro
Keyword(s):  
Vaccine Strain ◽  
Equine Infectious Anemia Virus ◽  
Genetically Engineered ◽  
Commercial Application ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Antibodies ◽  
Serological Method ◽  
Live Virus ◽  
Equine Infectious Anemia ◽  
Anemia Virus

ABSTRACT We recently reported a highly protective attenuated live virus vaccine for equine infectious anemia virus (EIAV) based on a proviral construct (EIAVUKΔS2) with a genetically engineered mutation in the viral S2 gene that eliminates expression of this accessory protein. While the EIAVUKΔS2 vaccine provides protection from detectable infection by experimental challenge with highly virulent virus, the potential for commercial application of this vaccine is complicated by the fact that horses inoculated with the EIAVUKΔS2 vaccine strain become seropositive in various reference diagnostic assays based on detection of antibodies to virion core or envelope proteins. To address this issue, we describe here the development and optimization of a new serologic EIAV diagnostic enzyme-linked immunosorbent assay (ELISA) to detect serum antibodies to the EIAV S2 protein that are produced in infected horses but not in horses inoculated with the EIAVUKΔS2 vaccine virus. The test S2 protein antigen was developed using the S2 gene sequence from the EIAVUK strain of virus and a series of modifications to facilitate production and purification of the diagnostic antigen, designated HS2G. Using this HS2G as antigen, we describe the development of an affinity ELISA that provides a sensitive and specific detection of S2-specific serum antibodies in experimentally and field-infected horses (22 of 24), without detectable reactivity with immune serum from uninfected (12 of 12) or vaccinated (29 of 29) horses. These data indicate that the S2-based diagnostic ELISA has the potential to accurately differentiate horses infected with EIAV from horses inoculated with an attenuated EIAV vaccine strain with a mutant S2 gene.

scholarly journals Surveillance of the equine infectious anemia virus in Eastern and Central Saudi Arabia during 2014-2016

2019 ◽  
Vol 12 (5) ◽  
pp. 719-723 ◽  
Author(s):  
Abdulmohsen Abdullah Alnaeem ◽  
Maged Gomaa Hemida
Keyword(s):  
Saudi Arabia ◽  
Equine Infectious Anemia Virus ◽  
Poor Performance ◽  
Enzyme Linked Immunosorbent Assay ◽  
Horse Population ◽  
Central Regions ◽  
Equine Infectious Anemia ◽  
Serological Surveillance ◽  
Anemia Virus ◽  
Equine Industry

Background: Equine infectious anemia virus (EIAV) is one of the most important threats to the equine industry globally. This is due to the poor performance of the affected horses, which requires euthanization of the infected animals upon the infection confirmation. Infected animals remain carriers throughout their life. EIAV infection has been reported in many parts of the world, including North America, Europe, Asia, and Africa. However, the EIAV status is never assessed in horses in the Gulf area, especially in the Kingdom of Saudi Arabia (KSA). Aim: This study aimed to perform molecular and serological surveillance among some horse populations in Eastern and Central Saudi Arabia. Materials and Methods: Sera and whole blood were collected from 361 horses and 19 donkeys from the eastern and central regions of Saudi Arabia during January 2014-December 2016. Sera were tested by the commercial EIAV enzyme-linked immunosorbent assay kits. Moreover, the collected blood samples were tested by the commercial real-time polymerase chain reaction kits. Results: Our serological surveillance revealed the absence of any antibodies against EIAV in the tested animals. Similar results were reported for the tested horses' plasma. This study confirms the absence of EIAV in horses and donkeys from Eastern and Central Saudi Arabia during the tenure of the current study. However, careful monitoring of the EIAV is highly recommended to avoid the emergence of such a virus in the horse population in Saudi Arabia. Conclusion: To the best of our knowledge, this is the first EIAV surveillance conducted not only in Saudi Arabia but also in the Gulf area. This study confirms the absence of EIAV in the tested equine population in the eastern and central regions of Saudi Arabia during 2014-2016.

scholarly journals Comparison of Diagnostic Tests for the Detection of Equine Infectious Anemia Antibody

1989 ◽  
Vol 1 (1) ◽  
pp. 50-52 ◽  
Author(s):  
Tatsuo Matsushita ◽  
Lyndal K. Hesterberg ◽  
James P. Porter ◽  
Barbara J. Smith ◽  
Louis E. Newman
Keyword(s):  
Western Blot ◽  
Diagnostic Tests ◽  
Equine Infectious Anemia Virus ◽  
Competitive Elisa ◽  
Enzyme Linked Immunosorbent Assay ◽  
Veterinary Services ◽  
Agar Gel ◽  
Equine Infectious Anemia ◽  
Low Levels ◽  
Anemia Virus

Two diagnostic tests are approved for detecting antibody to equine infectious anemia virus: the agar-gel immunodiffusion (AGID) test and the competitive enzyme-linked immunosorbent assay (ELISA). A total of 420 sera from National Veterinary Services Laboratories check sets were tested with the AGID and competitive ELISA. A 100% correlation was obtained. The AGID and competitive ELISA were further used to test difficult samples with low levels of equine infectious anemia antibody (weak positives). A third test (Western blot) was also used with these weak positive samples to resolve any discordant results.

scholarly journals Western Blot Assay Using Recombinant p26 Antigen for Detection of Equine Infectious Anemia Virus-Specific Antibodies

2007 ◽  
Vol 14 (12) ◽  
pp. 1646-1648 ◽  
Author(s):  
I. Alvarez ◽  
G. Gutierrez ◽  
E. Ostlund ◽  
M. Barrandeguy ◽  
K. Trono
Keyword(s):  
Western Blot ◽  
Capsid Protein ◽  
Diagnostic Tool ◽  
Equine Infectious Anemia Virus ◽  
Enzyme Linked Immunosorbent Assay ◽  
Agar Gel ◽  
Western Blot Assay ◽  
Specific Antibodies ◽  
Equine Infectious Anemia ◽  
Anemia Virus

ABSTRACT We analyzed the performance of a single-band Western blot (WB) test using recombinant p26 (rp26) capsid protein of equine infectious anemia virus. According to the results obtained, the rp26 WB test is a reliable confirmatory diagnostic tool to be used as a complementary test after an enzyme-linked immunosorbent assay or agar gel immunodiffusion test yielding doubtful results.

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