Enzyme-Linked Immunosorbent Assay (ELISA) is a widely used analytical technique in equine research for detecting and quantifying specific proteins, hormones, and antibodies in horse biological samples. This method relies on antigen-antibody interactions and employs enzyme-linked antibodies to produce a measurable signal, typically a color change, indicating the presence and concentration of the target molecule. ELISA is applicable in various areas of equine health, including the diagnosis of infectious diseases, monitoring of immune responses, and assessment of physiological conditions. It is valued for its specificity, sensitivity, and ability to process multiple samples simultaneously. This page compiles peer-reviewed research studies and scholarly articles that explore the applications, methodologies, and advancements of ELISA in equine science.
Huang J, Hartley CA, Ficorilli NP, Crabb BS, Studdert MJ.Glycoprotein G (gG) deletion mutants of EHV1 and EHV4, designated EHV1DeltagG and EHV4DeltagG, were constructed. The growth characteristics of the EHV1DeltagG mutants were similar to the parent virus. All of the EHV4DeltagG mutants grew more slowly in cell culture and produced plaques of different morphology including smaller size. The yields of both gG deletion mutant viruses in cell culture were similar to the parent viruses. Sequencing of the genes flanking gG, Southern blot, PCR and western blot analyses of the mutant viruses demonstrated that the deletions were as expected, except for EHV...
Hungerford NL, Sortais B, Smart CG, McKinney AR, Ridley DD, Stenhouse AM, Suann CJ, Munn KJ, Sillence MN, McLeod MD.Due to the potential for misuse of a wide range of anabolic steroids in horse racing, a screening test to detect multiple compounds, via a common class of metabolites, would be a valuable forensic tool. An enzyme-linked immunosorbent assay (ELISA) has been developed to detect 17alpha-alkyl anabolic steroid metabolites in equine urine. 16beta-Hydroxymestanolone (16beta,17beta-dihydroxy-17alpha-methyl-5alpha-androstan-3-one) was synthesised in six steps from commercially available epiandrosterone (3beta-hydroxy-5alpha-androstan-17-one). Polyclonal antibodies were raised in sheep, employing mesta...
Gingrich JB, Williams GM.Paucity of data on host-feeding patterns and behavior of 43 mosquito species that are reported as suspected West Nile virus (WN) vectors has limited full evaluation of their vectorial capacity. Recent studies addressing this issue need additional confirmation and should also be expanded to include collections of species or subpopulations attracted to humans. We used 4 types of collection methods to collect mosquitoes, including omnidirectional Fay-Prince traps, Centers for Disease Control-type light traps, gravid traps, and human-landing collections. Mosquitoes were collected during 2 full WN ...
Linklater WL, Henderson KM, Cameron EZ, Stafford KJ, Minot EO.To investigate the utility of faecal oestrone sulphate (OS) concentrations for detecting pregnancy in mares during behavioural studies of feral horses, in which the collection and preservation of samples is not immediate. Methods: Oestrone sulphate concentrations were measured in fresh dung samples collected from 153 free-roaming Kaimanawa mares throughout the year. In addition, multiple samples were taken from the same pile to investigate the reliability of diagnosis from a single sample, as well as the influence of time until preservation on OS concentrations. Samples were also taken before ...
Sofronic-Milosavljevic Lj, Ilic N, Djordjevic M, Savic M, Gruden-Movsesijan A, Cuperlovic K, Murrell KD.In the Balkan countries, where trichinellosis is a re-emerging zoonosis, it is of great importance to determine Trichinella infection prevalence among the major hosts, including horses. One method for monitoring prevalence is serological surveillance; however, the validity of serological methods in horses is not well understood. The dynamics of anti-Trichinella IgG production and circulating excretory/secretory (ES) antigens were investigated in three horses experimentally-infected with Trichinella spiralis. Horses were slaughtered at 32 week post infection (p.i.). Low worm burdens were found ...
Hoane JS, Yeargan MR, Stamper S, Saville WJ, Morrow JK, Lindsay DS, Howe DK.Neospora hughesi is a recently identified cause of equine protozoal myeloencephalitis. However, the significance of this parasite is poorly understood. An enzyme-linked immunosorbent assay (ELISA) with a recombinant form of the N. hughesi 29-kDa surface antigen (rNhSAG1) was developed for serodiagnosis of equine N. hughesi infections. Parallel ELISA analysis showed that animals immunized or infected with N. hughesi exhibited greater antibody reactivity with rNhSAG1 than with the Neospora caninum homolog, rNcSAG1. The rNhSAG1 ELISA showed 94.4% sensitivity and 95.0% specificity when compared wi...
Li F, Stevenson RA, Crabb BS, Studdert MJ, Hartley CA.Equine rhinitis A virus (ERAV) is a significant pathogen of horses and is also closely related to Foot-and-mouth disease virus (FMDV). Despite these facts, knowledge of the prevalence and importance of ERAV infections remains limited, largely due to the absence of a simple, robust diagnostic assay. In this study, we compared the antigenicities of recombinant full-length and fragmented ERAV capsid proteins expressed in Escherichia coli by using sera from experimentally infected and naturally exposed horses. We found that, from the range of antigens tested, recombinant proteins encompassing the ...
Hartley CA, Wilks CR, Studdert MJ, Gilkerson JR.To compare methods of detecting equine herpesvirus type 1 (EHV1)- and EHV4-specific antibodies in horse sera. Methods: 33 acute and convalescent serum samples from experimentally or naturally infected horses after confirmed EHV1 or EHV4 infection. Methods: For each sample, serum antibody titers against EHV1 and EHV4 were determined by use of virus neutralization (VN) and complement fixation (CF) assays. The ELISA absorbance values for each serum sample were determined against the EHV1 and EHV4 recombinant ELISA antigens. Values obtained for acute and convalescent sera in each assay were compar...
Wagner B, Robeson J, McCracken M, Wattrang E, Antczak DF.Recombinant cytokines are valuable tools for functional studies and candidates for vaccine additives or therapeutic use in various diseases. They can also be used to generate specific antibodies to analyze the roles of different cytokines during immune responses. We generated a mammalian expression system for recombinant cytokines using the equine IgG1 heavy chain constant region as a tag for detection and purification of the expressed cytokine, demonstrated here using equine interferon-gamma (IFN-gamma), interleukin-2 (IL-2), interleukin-4 (IL4) and transforming growth factor-beta1 (TGF-beta1...
Nieto JE, Aldridge BM, Beldomenico PM, Aleman M, Snyder JR.To determine the nucleotide sequence of the equine intestinal fatty acid binding protein (I-FABP) gene, its expression in various regions of the gastrointestinal tract, and the use of measuring I-FABP in horses with colic. Animals-86 horses with colic. Methods: The mRNA sequence for the I-FABP gene was obtained by use of a rapid amplification of complementary DNA ends technique. Comparative I-FABP gene expression was quantitated by use of a real-time reverse transcription-polymerase chain reaction assay. Amounts of I-FABP in abdominal fluid and plasma were measured by use of an ELISA kit. Asso...
Magnarelli L, Fikrig E.Blood samples were collected from 98 horses suspected of having borreliosis or granulocytic ehrlichiosis in Connecticut and New York State, USA during 1985, 1995, and 1996. Serum antibodies to Borrelia burgdorferi were detected by an enzyme-linked immunosorbent assay (ELISA), based on whole-cell and recombinant antigens, in 82 (84%) horses. Of the 181 sera tested, 59% were positive, using whole-cell antigens, compared to 48% with protein (p)37 and 35% with VlsE antigens. An ELISA containing either of these fusion proteins can be used as an adjunct to general screening by an ELISA or immunoblot...
Nishita T, Takahasi M, Kasuya T, Matsui K, Ichihara N, Murakami M, Asari M.Equine carbonic anhydrase isozymes (CA-I and CA-II) were purified from erythrocytes by several column chromatography. Polyclonal anti-CA-I and anti-CA-II sera were produced in rabbits. Sensitive competitive enzyme-linked immunosorbent assays (ELISA) were established to determine the developmental changes in CA-I and CA-II levels in equine erythrocytes. Concentrations of CA-I and CA-II in erythrocytes from 150 clinically normal thoroughbreds (123 racehorses and 27 riding horses) were determined by ELISA. Mean (+/- SD) concentrations of CA-I and CA-II in racehorses were 1.70 +/- 0.48 and 0.94 +/...
Hodgkinson JE, Freeman KL, Lichtenfels JR, Palfreman S, Love S, Matthews JB.The efficacy of five daily fenbendazole (FBZ) treatments was tested against benzimidazole-resistant cyathostomins in naturally infected horses (n=13). Horses were treated with pyrantel embonate (PYR) to remove adult strongyles followed, 7 days later, by a 5-day course of FBZ. The PYR treatment produced an average faecal egg count reduction of 98%. All samples were negative by faecal egg count 7 days after the start of the FBZ treatment. Positive egg counts were observed from 28 days after the start of FBZ treatment and all horses displayed positive faecal egg counts by 77 days after treatment....
Gutmann S, Zawatzky R, Müller M.Interferon-gamma (IFN-gamma) is a key cytokine in cell-mediated immunity. To measure IFN-gamma production of equine lymphocytes (eqIFN-gamma), we developed a quantitative ELISA. Monoclonal antibodies (mAb) were produced against bacterially derived eqIFN-gamma. The mAbs recognised recombinant and lymphocyte-derived eqIFN-gamma in ELISA, Western blotting, as well as flow cytometric and microscopic analysis. In contrast to bacterially derived material, mammalian and insect cell-derived eqIFN-gamma was biologically active but could be neutralised by one of the monoclonal antibodies. Unexpectedly, ...
Maree S, Paweska JT.This paper describes the production and purification of a group-specific recombinant protein VP7 of African horse sickness virus serotype 3 (AHSV-3) and validation of an I-ELISA for the detection of IgG-antibodies to VP7 in horse sera. Baculovirus-expressed VP7 crystals were purified from infected insect cells. Analytical accuracy of the I-ELISA was examined using sera (n = 38) from an experimentally infected horse, from foals born to vaccinated mares, from guinea-pigs immunized with nine serotypes of AHSV, and from sera of animals infected with other orbiviruses. Compared to traditional serol...
Maeda K, Mizukoshi F, Hamano M, Kai K, Kondo T, Matsumura T.Recently, a novel 12-mer B-cell epitope, MKNNPIYSEGSL, in the type-specific region of equine herpesvirus 1 (EHV-1) glycoprotein G (gG) was identified and used as an antigen for enzyme-linked immunosorbent assay (Maeda et al., J. Clin. Microbiol. 42:1095-1098, 2004). Although our prototype strain, TH20p, possesses two repeat sequences containing the B-cell epitope, the EHV-4 NS80567 strain has two repeat sequences that are not identical. One repeat sequence stretch contained the B-cell epitope, while the other contained the 11-mer, MKNNPVYSESL (underlining indicates a different amino acid). In ...
Porter MB, Long M, Gosche DG, Schott HM, Hines MT, Rossano M, Sellon DC.The West Nile (WN) virus, present in the United States since 1999, is a cause of encephalomyelitis in birds, alligators, humans, and horses. No data exist regarding detection of anti-WN virus immunoglobins in equine cerebrospinal fluid (CSF). The aims of this study were to evaluate the blood-brain barrier (BBB) in WN virus-infected (WNE) horses, to compare diagnostic testing in serum and CSF, and to describe the immunoglobulin M (IgM) response in serum and CSF of vaccinated horses. CSF was collected from the lumbosacral (LS) space (n = 13) or the allanto-occipital (AO) space (n = 14) of WNE ho...
Boldbaatar D, Xuan X, Battsetseg B, Igarashi I, Battur B, Batsukh Z, Bayambaa B, Fujisaki K.The purpose of this study was to demonstrate the occurrence of equine piroplasmosis in Mongolia, a country in which the disease occurs epidemically in different climatic conditions. Antibodies to Babesia equi and B. caballi were determined in serum samples of 254 pastured horses in different locations of Mongolia using an enzyme-linked immunosorbent assay with recombinant antigens. One hundred and eighty-five (72.8%) and 102 (40.1%) of all serum samples were positive for B. equi and B. caballi infections, respectively. In addition, 78 (30.7%) samples were positive for both B. equi and B. cabal...
Skiöldebrand E, Heinegård D, Eloranta ML, Nilsson G, Dudhia J, Sandgren B, Ekman S.The aim of the present study was to correlate the levels of COMP and aggrecan as indicators of tissue damage, in synovial fluid (sf) from carpal joints of acutely lame racehorses, with macroscopical lesions of articular cartilage (OA), osteochondral fractures and ligament tears found at arthroscopy. Sixty-three lame horses [49 Standardbred trotters (STB) and 14 Thoroughbreds (TB)] in conventional training and racing that underwent arthroscopy of their middle carpal or radiocarpal joints were included in the study. Intact as well as fragmented COMP and aggrecan released into the synovial fluid ...
Paré J, Simard C.The purpose of this study was to estimate the performance characteristics (accuracy, detection limit, and precision) of commercially available enzyme-linked immunosorbent assay (ELISA) and agar gel immunodiffusion (AGID) kits in comparison with a reference AGID kit for the detection of equine infectious anemia (EIA) antibodies in horses for regulatory use in Canada. A total of 285 positive and 315 negative samples by the reference AGID were tested blindly on 2 other AGID and 4 ELISA kits. Commercially available AGID kits for the serodiagnosis of EIA were found equivalent. The 3 ELISAs directed...
Kania SA, Reinemeyer CR.Anoplocephala perfoliata has a prevalence as high as 60% in some geographical areas and has been associated with increased risk for serious conditions in the horse including bowel irritation, ileal impaction, and spasmodic colic. Identification of infected animals based upon detection of eggs in feces is labor intensive and unreliable. This study involved the development of a test for A.perfoliata coproantigen using an antigen capture enzyme linked immunosorbent assay (ELISA) and correctly distinguished between infected and uninfected animals in a trial with a small sample size.
Jin S, Issel CJ, Montelaro RC.We recently reported a highly protective attenuated live virus vaccine for equine infectious anemia virus (EIAV) based on a proviral construct (EIAVUKDeltaS2) with a genetically engineered mutation in the viral S2 gene that eliminates expression of this accessory protein. While the EIAVUKDeltaS2 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 EIAVUKDeltaS2 vaccine strain become seropositive in various reference diagnosti...
Konishi E, Shoda M, Ajiro N, Kondo T.Antibodies to Japanese encephalitis virus (JEV) nonstructural 1 (NS1) protein constitute a marker of natural JEV infection among populations vaccinated with inactivated JE vaccine. In Japan, with few recent human JE cases, the natural infection rate is critical to evaluate the necessity of continuing JE vaccination. A sensitive immunochemical staining method for detecting NS1 antibodies in individuals naturally and subclinically infected with JEV was previously established. Here, an enzyme-linked immunosorbent assay (ELISA) to detect NS1 antibodies in equine sera was developed and evaluated as...
Arroyo LG, Weese JS, Staempfli HR.Despite empirical clinical association of infection with Clostridium difficile with colitis in horses, a causal link has not been confirmed. The objective of this study was to develop a model of C. difficile-associated diarrhea in foals with normal transfer of passive immunity. Nine 1-day-old pony foals were inoculated intragastrically with spores or vegetative cells of C. difficile. Five foals were challenged with spores, with 2 receiving 10(5) colony-forming units (CFUs) and concurrently 3 receiving 10(7) CFUs once daily for 3 days. Clindamycin was administered orally to disrupt gastrointest...
Popot MA, Bonnaire Y, Guéchot J, Toutain PL.Hyaluronic acid (HA) is an endogenous glycosaminoglycan used in the treatment of joint diseases, but medication control is required by horseracing authorities. Therefore, a medication control policy needs to be established. Objective: To establish physiological plasma HA concentrations in post race horses, determine the HA endogenous production rate and document the disposition of HA after i.v. and intra-articular hyaluronic acid administration at recommended therapeutic doses. Methods: Hyaluronan concentrations in plasma were determined using an ELISA specific test; concentrations in synovial...
Singh BK, Ahuja S, Gulati BR.A single-dilution, sensitive and specific monoclonal antibody-based blocking enzyme-linked immunosorbent assay (B-ELISA) was developed as an alternative to the cumbersome virus neutralization test (VNT) for detection of equine herpesvirus-1 (EHV-1) antibodies. Neutralizing monoclonal antibodies (1H6 and 9C6) raised against EHV-1 (Hisar-90-7 strain) and sera from 70 horses (30 known negative and 40 known positive for EHV-1 antibodies by VNT) were used for standardization of the B-ELISA. Using a single serum dilution of 1:250 in B-ELISA, 100% specificity was obtained with both monoclonal antibod...
Quirin R, Salas M, Zientara S, Zeller H, Labie J, Murri S, Lefrançois T, Petitclerc M, Martinez D.To determine whether West Nile virus (WNV) had reached the archipelago of Guadeloupe, a serologic study in horses and birds was conducted in 2002. Immunoglobulin (Ig) G, IgM, enzyme-linked immunosorbent assay, and seroneutralization tests identified WNV infection in horses and chickens. Six months later, a high rate of seroconversion was observed in horses.
Lehner AF, Hughes CG, Harkins JD, Nickerson C, Mollett B, Dirikolu L, Bosken J, Camargo F, Boyles J, Troppmann A, Karpiesiuk WW, Woods WE, Tobin T.We have investigated the detection, confirmation, and metabolism of the beta-adrenergic agonist ractopamine administered as Paylean to the horse. A Testing Components Corporation enzyme-linked imunosorbent assay (ELISA) kit for ractopamine displayed linear response between 1.0 and 100 ng/mL with an I-50 of 10 ng/mL and an effective screening limit of detection of 50 ng/mL. The kit was readily able to detect ractopamine equivalents in unhydrolyzed urine up to 24 h following a 300-mg oral dose. Gas chromatography-mass spectrometry (GC-MS) confirmation comprised glucuronidase treatment, solid-pha...
Foote CE, Love DN, Gilkerson JR, Whalley JM.A silent cycle of equine herpesvirus 1 infection has been described following epidemiological studies in unvaccinated mares and foals. In 1997, an inactivated whole virus EHV-1 and EHV-4 vaccine was released commercially in Australia and used on many stud farms. However, it was not known what effect vaccination might have on the cycle of infection of EHV-1. Objective: To investigate whether EHV-1 and EHV-4 could be detected in young foals from vaccinated mares. Methods: Nasal and blood samples were tested by PCR and ELISA after collection from 237 unvaccinated, unweaned foals and vaccinated an...
Barrett EJ, Farlam J, Proudman CJ.Two hundred and thirty-three horses were screened for the presence of roundworms by faecal egg counts (FECs) and for tapeworms by an ELISA specific for antibodies to the immunodominant 12 kDa and 13 kDa tapeworms antigen. The 62 horses were found to be infected with both parasites were treated with a combination of 0.2 mg/kg ivermectin and 1.5 mg/kg praziquantel. The treatment suppressed the median FEC of the horses to zero for 10 weeks and significantly reduced their anti-12/13 kDa antibody levels. The estimated risk of tapeworm-associated colic in these horses was halved by 12 weeks after th...
Lee DH, Lee EB, Seo JP, Ko EJ.Despite vaccination, equine influenza virus (EIV) and equine herpesvirus (EHV) infections still cause highly contagious respiratory diseases in horses. Recently, concurrent vaccination with EIV and EHV was suggested as a new approach; however, there have been no reports of concurrent vaccination with recombinant canarypox EIV and inactivated EHV vaccines. In this study, we aimed to compare the EIV-specific immune responses induced by concurrent administrations of a recombinant canarypox EIV vaccine and an inactivated bivalent EHV vaccine with those induced by a single recombinant canarypox EIV...
Horsington JJ, Gilkerson JR, Hartley CA.Erbovirus is a genus of the family Picornaviridae and equine rhinitis B virus (ERBV) is the sole species. Erboviruses infect horses causing acute respiratory disease and sub-clinical and persistent infections. Despite the high seroprevalence and worldwide distribution of these viruses, the pathogenesis and antigenic structure of the three ERBV serotypes (ERBV1, 2 and 3) is poorly understood. To characterise linear epitopes on ERBV structural proteins, a set of fusion proteins were expressed in Escherichia coli. These proteins were tested in Western blot and ELISA and reactive proteins were als...
Mayo C, Venter E, Steyn J, Coetzee P, van Vuuren M, Crafford J, Schütte C, Venter G.The seasonal abundance of Culicoides midges, the vector of Bluetongue and African horse sickness viruses (BTV/AHSV) and the presence of viruses in midges were determined in 3 geographic areas in South Africa. In the Onderstepoort area, more than 500,000 Culicoides midges belonging to 27 species were collected. Eighteen midge species were collected throughout Winter and the presence of AHSV and BTV RNA in midges was detected using real time reverse transcription quantitative polymerase chain reaction. The nucleic acid of AHSV was found in 12 pools out of total pools of 35 Culicoides. Twenty‑f...
Kitai Y, Shirafuji H, Kanehira K, Kamio T, Kondo T, Konishi E.West Nile virus (WNV) and Japanese encephalitis (JE) virus are distributed separately in the world with some exceptions. There is a concern that WNV may invade into Asia where JE virus exists. On and after such invasion, any differential diagnosis could be complicated by serological crossreactivities. We previously demonstrated experimentally using horses infected with WNV that preimmunization with inactivated JE vaccine considerably affected the ability of neutralization tests and immunoglobulin M (IgM) antibody-capture enzyme-linked immunosorbent assay (ELISA) to diagnose WNV infection. Here...
Keller LE, Tait Wojno ED, Begum L, Fortier LA.Infiltration of cluster of differentiation (CD) 3 (CD3) T cells into the synovium and synovial fluid occurs in most patients with posttraumatic osteoarthritis. During disease progression, proinflammatory T helper 17 cells and anti-inflammatory regulatory T cells infiltrate the joint in response to inflammation. This study aimed to characterize the dynamics of regulatory T and T helper 17 cell populations in synovial fluid from equine clinical patients with posttraumatic osteoarthritis to determine whether phenotype and function are associated with potential immunotherapeutic targets. An imbala...
Xuan X, Igarashi I, Tanaka T, Fukumoto S, Nagasawa H, Fujisaki K, Mikami T.A latex agglutination test (LAT) using recombinant equi merozoite antigen 1 (EMA-1) for the detection of antibodies to Babesia equi was developed. The LAT was able to differentiate very clearly between sera from B. equi-infected horses and sera from Babesia caballi-infected horses or from normal horses. The LAT results were identical to those of a previously developed enzyme-linked immunosorbent assay. These results indicate that LAT using recombinant EMA-1 might be very useful as a routine screening method for the diagnosis of B. equi infection.
Jenvey C, Caraguel C, Howarth GB, Riley CB.Prior to the start of endogenous production of immunoglobulins (Igs), absorption of maternal Igs is important to protect against pathogens in the early neonatal period. It is possible that mare- or foal-associated factors may influence neonatal IgA concentrations. Objective: The temporal relationships among serum and milk IgA concentrations in Thoroughbred mare-foal pairs were explored to determine if periparturient mare- and foal-associated factors contribute to the prediction of foal serum IgA concentrations. Methods: Blood and milk samples as well as complete veterinary records, were collec...
Intan-Shameha AR, Divers TJ, Morrow JK, Graves A, Olsen E, Johnson AL, Mohammed HO.The current study aimed at the investigating the potential use of phosphorylated neurofilament H (pNF-H) as a diagnostic biomarker for neurologic disorders in the horse. Paired serum and cerebrospinal fluid (CSF) samples (n=88) and serum only (n=30) were obtained from horses diagnosed with neurologic disorders and clinically healthy horses as control. The neurologic horses consisted of equine protozoal myeloencephalitis (EPM) (38 cases) and cervical vertebral malformation (CVM) (23 cases). Levels of pNF-H were determined using an ELISA. The correlation between CSF and serum concentrations of p...
Okumura M, Fujinaga T.To establish a sensitive and specific monoclonal antibody (MAB) against equine keratan sulfate (KS) and to develop an enzyme immunoassay for measurement of the concentration of KS in serum and synovial fluid from horses. Methods: 18 synovial fluid and 48 serum samples were obtained from clinically normal horses and horses with arthritis. Methods: BALB/c mice were immunized with chondroitinase-ABC-digested proteoglycan monomer from equine joint cartilage, and MAB were raised, using Sp2/O cells as a fusion partner. A competitive ELISA was optimized, using one of the established MAB, and KS conce...
du Plessis DH, van Wyngaardt W, Bremer CW.African horsesickness virus (AHSV), an important disease of equines is caused by an orbivirus. Because of the need to contain the spread of the disease, it is often essential to make a rapid diagnosis. For this purpose, an ELISA capable of detecting viral antigen in animal tissue and in cell culture fluid was developed. Immobilised F(ab')2 fragments prepared by digestion of AHSV-specific IgG with pepsin were used to trap virus from tissue homogenates or cell culture supernatant. After addition of intact IgG as detecting antibody, Staphylococcus aureus protein A labelled with horseradish peroxi...
Toth B, Slovis NM, Constable PD, Taylor SD.Bacterial sepsis remains a leading cause of morbidity and mortality in neonatal foals, but accurate diagnostic and prognostic markers are lacking. Adrenomedullin (AM) is a polypeptide with diverse biologic effects on the cardiovascular system that increases in septic humans and laboratory animals. Objective: Plasma AM concentration (p[AM]) is increased in septic neonatal foals compared to sick nonseptic and healthy control foals, and p[AM] is predictive of survival in septic neonatal foals. Methods: Ninety critically ill (42 septic, 48 sick nonseptic) and 61 healthy foals <1 week of age. Metho...
Taesuji M, Rattanamas K, Kulthonggate U, Mamom T, Ruenphet S.The immune responses of animals infected with African horse sickness (AHS) virus are determined by enzyme-linked immunosorbent assay (ELISA), complement fixation, and virus neutralization test. During the outbreaks of AHS in Thailand, the immune response after vaccination has been monitored using commercial test kits such as blocking ELISA, which are expensive imported products unavailable commercially in Thailand. This study aimed to assess the sensitivity and specificity of anti-AHS virus antibodies using dot blotting based on monovalent and polyvalent strains of live attenuated AHS vaccine....
Yarnell K, Purcell RS, Walker SL.Adrenal activity can be assessed in the equine species by analysis of feces for corticosterone metabolites. During a potentially aversive situation, corticotrophin releasing hormone (CRH) is released from the hypothalamus in the brain. This stimulates the release of adrenocorticotrophic hormone (ACTH) from the pituitary gland, which in turn stimulates release of glucocorticoids from the adrenal gland. In horses the glucocorticoid corticosterone is responsible for several adaptations needed to support equine flight behaviour and subsequent removal from the aversive situation. Corticosterone met...
Getachew AM, Innocent G, Proudman CJ, Trawford A, Feseha G, Reid SW, Faith B, Love S.A 12/13 kDa antigen, tapeworm ELISA test, developed for use in horses, was used to detect parasite-specific serum antibody, IgG(T), in the serum of donkeys. In a pilot study the 12/13 kDa antigen was tested and proved to detect the antibody, IgG(T), in donkey sera. Blood samples from 797 donkeys, naturally exposed to cestode infection, from four geographical localities were collected and sera were prepared and analysed. There was substantial serological evidence that donkeys were potentially infected with A. perfoliata. A range of ELISA OD values were obtained from the serological assay. Over ...
Arias M, Yeargan M, Francisco I, Dangoudoubiyam S, Becerra P, Francisco R, Sánchez-Andrade R, Paz-Silva A, Howe DK.Horses serve as an intermediate host for several species of Sarcocystis, all of which utilize canids as the definitive host. Sarcocystis spp. infection and formation of latent sarcocysts in horses often appears to be subclinical, but morbidity can occur, especially when the parasite burden is large. A serological survey was conducted to determine the presence of antibodies against Sarcocystis spp. in seemingly healthy horses from the Galicia region of Spain. Western blot analyses using Sarcocystis neurona merozoites as heterologous antigen suggested greater than 80% seroprevalance of Sarcocyst...
Sams RA, Gerken DF, Ashcraft SM.The objectives of the study were to compare various methods to determine flunixin in test samples collected periodically from horses after intramuscular (IM) and intravenous (IV) dosing at the maximum recommended dosage and to document detection times for this drug in test samples. Flunixin, a nonsteroidal anti-inflammatory drug approved for use in horses, was administered to eight mares in five consecutive daily doses of 1.1 mg per kilogram of body weight by the IM or IV route. Flunixin was detected in urine samples collected at various times after drug administration by flunixin enzyme-linke...
Topper MJ, Prasse KW.To evaluate new ELISA for measurement of thrombin-antithrombin III (TAT) concentration, and to correlate the values to other tests of hemostasis in horses with colic. Methods: Plasma TAT concentration and 8 other hemostasis analytes were measured in horses with colic at hospital admission and during the next 4 days. Retrospectively, data were analyzed by outcome, broad-category diagnosis, and clinical management, and for correlation between TAT and other assays. Methods: 100 horses with colic. Methods: Plasma samples were evaluated for TAT, fibrinogen, and fibrin degradation products concentra...
Hamblin C, Mellor PS, Graham SD, Hooghuis H, Montejano RC, Cubillo MA, Boned J.A total of 256 sera collected from three species of domesticated equidae in four different Spanish provinces were examined 1-4 months after the administration of attenuated monovalent African horse sickness virus (AHSV) serotype 4 vaccine. Approximately 10% of the sera were negative by ELISA, virus neutralization, agar gel immuno-diffusion and complement fixation tests. Similar negative reactions were recorded with sera from two ponies after experimental primary vaccination. The rapid rise in antibodies in sera from these two ponies, after a second dose of vaccine, suggested they would probabl...
Singha H, Goyal SK, Malik P, Khurana SK, Singh RK.Equine infectious anemia (EIA)-a retroviral disease caused by equine infectious anemia virus (EIAV)-is a chronic, debilitating disease of horses, mules, and donkeys. EIAV infection has been reported worldwide and is recognized as pathogen of significant economic importance to the horse industry. This disease falls under regulatory control program in many countries including India. Control of EIA is based on identification of inapparent carriers by detection of antibodies to EIAV in serologic tests and "Stamping Out" policy. The current internationally accepted test for diagnosis of EIA is the ...
Perryman LE, O'Rourke KI, Mason PH, McGuire TC.Equine-murine xenohybridoma cells were produced using SP2/0 murine myeloma cells and splenic lymph node cells obtained from horses infected with 10(6) TCID50 of single cloned variants of equine infectious anaemia virus (EIAV). The xenohybridomas secreted equine IgG monoclonal antibodies reactive with EIAV in enzyme immunoassays employing purified virus. Seven antibodies were studied in detail. They bound to viral glycoproteins (gp90 or gp45) in radioimmunoprecipitation assays, and reacted with homologous EIAV as well as five other cloned variants of EIAV. When evaluated against a single cloned...
Thomas LM, Huntington PJ, Mead LJ, Wingate DL, Rogerson BA, Lew AM.The use of the bacterial expression vector, pGex, to produce an abundant, soluble fusion protein of gp45 from equine infectious anaemia virus is described. Purification of the recombinant protein was achieved by one step affinity chromatography on immobilized glutathione using competitive elution so no harsh conditions were required. This provides a readily available antigen that is defined, plentiful and cheap. Yields of 3.5 mg of purified soluble protein/litre of bacterial culture were obtained. This antigen was found to be suitable for ELISA. Background reactivity to either the glutathione-...
Saad B, Corradin G, Bosshard HR.The antigenic determinants for two monoclonal antibodies directed against horse apo-cytochrome c, a protein of disordered structure, as judged by spectroscopic and hydrodynamic criteria, have been studied by a combination of methods: antigen competition in solution by radio immunoassay and enzyme-linked immunoassay, and differential acetylation of free and antibody-bound antigen. In the latter method the accessibility of lysine residues of the antigen in the antigen-antibody complex is compared to the accessibility in the free antigen. The two antibodies against the heme-free protein do not re...
Gáspárdy A, Gallagher G, Bartha B, Cseh S, Fekete SG, Somoskői B.The authors aimed to determine the plasma melatonin concentration in mares and their new-born foals in the early post-partum period. Blood samples were collected from the jugular vein of 53 mare-foal pairs within twelve hours after parturition. Plasma melatonin levels were measured by ELISA. The melatonin concentration, adjusted for the moment of parturition using a generalised linear model, was 34.58 pg mL-1 in mares. It was significantly lower (27.63 pg mL-1) in the new-born foals. However, the melatonin concentration declined differently by the end of the twelve hours, it decreased less in...
Holbrook TC, McFarlane D, Schott HC.The inflammatory and neuroendocrine response to endurance exercise and relationship of these parameters to performance is not well documented in horses. HYPOTHESES OR OBJECTIVES: Evidence of systemic inflammation is associated with poor performance in horses competing in endurance events. Methods: Blood was collected prior to and at the finish or elimination point from horses competing in both the 80 and 160 km American Endurance Ride National Championship competitions in 2006. Immunoreactive alpha-melanocyte stimulating hormone (α-MSH) and tumour necrosis factor-alpha (TNF-α) were quantifie...
Yang G, Zhou B, Chen K, Hu Z, Guo W, Wang X, Du C. () and () are the causative pathogens of Equine piroplasmosis (EP), a disease that has brought huge economic losses and great restrictions to the global equine industry. Rapid and accurate diagnostic methods are critical for the effective monitoring of the disease. In this study, we developed novel competitive ELISA methods and western blot assays based on the EMA1 or Bc48 proteins to detect antibodies against or , respectively. In the novel cELISA, horseradish peroxidase (HRP)-labeled monoclonal antibodies are used in place of enzyme-conjugated secondary antibodies, in order to speed up th...
Okumura M, Kim GH, Tagami M, Haramaki S, Fujinaga T.Keratan sulphate (KS) concentration in sera from resting horses and horses training daily on a racetrack was measured by an inhibition enzyme-linked immunosorbent assay (ELISA) using anti-equine KS antibody 1/14/16H9. For the in-training horses, serum KS concentrations in 2-year-old-horses was significantly higher than 3- or 4-year-old-horses. A higher concentration of serum KS was found in the in-training group than in the long-term resting group in 2-year-old-horses. Serum KS concentration increased remarkably immediately after training in healthy horses, and at 1, 5, 9 and 24 h after traini...
Frey J, Eberle S, Stahl C, Mazuet C, Popoff M, Schatzmann E, Gerber V, Dungu B, Straub R.In Europe the incidence of botulism in horses has increased in the last decade due to the growing popularity of haylage feeding. Recombinant vaccines are safer and less expensive to produce and are generally better tolerated than toxoids. Objective: To investigate whether the recombinant C-terminal half of the heavy chain of the botulinum neurotoxin C (Hc BoNT/C) in combination with an immunstimulatory adjuvant is an appropriate vaccine candidate for horses by testing its efficacy to induce neutralising antibodies and by comparing its immunogenic properties and adverse reactions to a commercia...
Kummer LL, Govaere J, Egri B.Twenty-eight warmblood mares were monitored during their late pregnancy in the Teaching Hospital of Ghent University. The reliability of two commercial assays (enzyme immunoassay and glutaraldehyde coagulation test) used for determining the IgG concentrations of their newborn foals was tested. Mammary secretions were examined at the time of foaling (T0), and then 4 (T1) and 8 (T2) hours after foaling by refractometry and electrophoresis. The foals' blood IgG levels were measured at T1 and T2 as a routine clinical diagnostic examination using two different commercial test kits (SNAP Foal Ig and...
Rissanen K, Galat M, Kovalenko G, Rodnina O, Mikharovskyi G, Must K, Jokelainen P.Horses are recognized as important hosts for the zoonotic parasite Toxoplasma gondii, mainly because meat of infected horses can be a source of human T. gondii infections. However, the baseline knowledge on equine T. gondii infections is limited and lacking from many countries. This seroepidemiological study was set in Ukraine, a country where little is known about T. gondii prevalence in any host species. Methods: We investigated 78 serum samples from horses, collected from Kyiv and Lviv regions of Ukraine, using a commercial enzyme-linked immunosorbent assay (ELISA) and a commercial modified...
Harkins JD, Mundy GD, Woods WE, Lehner A, Karpiesiuk W, Rees WA, Dirikolu L, Bass S, Carter WG, Boyles J, Tobin T.Lidocaine is a local anaesthetic agent that is widely used in equine medicine. It is also an Association of Racing Commissioners International (ARCI) Class 2 foreign substance that may cause regulators to impose substantial penalties if residues are identified in post race urine samples. Therefore, an analytical/pharmacological database was developed for this drug. Using our abaxial sesamoid local anaesthetic model, the highest no-effect dose (HNED) for the local anaesthetic effect of lidocaine was determined to be 4 mg. Using enzyme-linked immunosorbent assay (ELISA) screening, administration...
