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.
Knowles DP, Kappmeyer LS, Stiller D, Hennager SG, Perryman LE.Horses infected with Babesia equi were previously identified by the presence of antibodies reactive with a merozoite surface protein epitope (D. P. Knowles, Jr., L. E. Perryman, L. S. Kappmeyer, and S. G. Hennager. J. Clin. Microbiol. 29:2056-2058, 1991). The antibodies were detected in a competitive inhibition enzyme-linked immunosorbent assay (CI ELISA) by using monoclonal antibody 36/133.97, which defines a protein epitope on the merozoite surface. The gene encoding this B. equi merozoite epitope was cloned and expressed in Escherichia coli. The recombinant merozoite protein, designated equ...
Su X, Morris DD, Crowe NA, Moore JN, Fischer KJ, McGraw RA.We describe the production and purification of recombinant equine tumor necrosis factor alpha (rETNF alpha), generation and characterization of murine monoclonal antibodies (Mabs) and rabbit polyclonal antibodies (Pabs) against ETNF alpha, and development of a sensitive enzyme-linked immunosorbent assay (ELISA). Genomic-derived DNA sequences encoding mature ETNF alpha were reconstructed by the polymerase chain reaction (PCR) and oligonucleotide-directed mutagenesis and were cloned into the vector pFLAG-1 for expression in Escherichia coli. rETNF alpha was purified by anti-FLAG immunoaffinity c...
Bürki F, Rossmanith W, Rossmanith E.Serological diagnosis of equine infectious anemia is of necessity group-reactive, i.e. based on viral core protein p26, because viral envelope components as well as the host's immune response to them undergo rapid antigenic change. Since 1970 the agar gel-immunodiffusion test ("Coggins-test") has been the diagnostic method of choice. Recently, ELISA tests have been introduced for faster and theoretically more sensitive serodiagnosis, while Western blots have been used to clarify doubtful results obtained in Coggins-tests. A commercial competitive ELISA was found to give practically equivalent ...
Cohen ND, Heck FC, Heim B, Flad DM, Bosler EM, Cohen D.Four hundred sixty-nine serum samples were obtained from horses admitted to the internal medicine service of the Texas Veterinary Medical Center between Jan 1 and Dec 31, 1990. Serum samples were tested by ELISA for antibody to Borrelia burgdorferi. Of these 469 samples, 1 (0.2%) was repeatedly seropositive for the organism by ELISA. Confirmatory testing by protein immunoblot was negative. The observed seroprevalence was 0%; the upper bound of the 95% confidence interval was 0.6%. These findings indicate the evidence of infection with B burgdorferi is presently uncommon in horses in central Te...
Sinclair R, Mumford JA.An antigen capture enzyme-linked immunosorbent assay (ELISA) was developed for the detection of equine herpesvirus type-1 (EHV-1) antigens in nasal swab specimens. The test was designed as a solid phase, amplified sandwich assay in which an EHV-1 specific monoclonal antibody was used to capture virus antigen and polyclonal antisera used to detect antigen bound to the test plates. Eight monoclonal antibodies were tested for their ability to capture virus antigen and one was selected for routine use. The sensitivity and specificity of the ELISA was compared with that of virus isolation using swa...
O'Brien MA, Holmes MA, Duffus WP.Anti-tetanus toxoid (TT) antibody (Ig) levels in the supernatant of cultured, pre-immunised equine peripheral blood mononuclear cells (PBMC) were measured by an indirect enzyme-linked immunoabsorbent assay (ELISA). Optimal anti-TT Ig production occurred at concentrations of stimulating, purified TT of between 0.001 and 0.1 micrograms ml-1, which varied depending on the cell concentration. Optimal anti-TT Ig production was most consistently produced when the cell concentration was 5 x 10(6) ml-1. At this cell concentration maximal anti-TT Ig was induced using 0.1 micrograms ml-1 TT. At a cell c...
Sanada Y, Noda H, Nagahata H.Serological survey of Rhodococcus equi infection in horses in Hokkaido was performed using ELISA. Of 2,879 horse sera, 318 (11.0%) gave antibody-positive (OD greater than or equal to 0.3) reactions. The antibody-positive rate of female was significantly higher (p less than 0.01) than that of male, and no statistical difference between Anglo-Arab and thoroughbred was detected in the antibody-positive rate. The maximum antibody-positive rate (27.1%) was shown at 14 years of age. The antibody-positive rates on the 160 farms were found to vary widely from 0 to 78.9%. A significant difference (p le...
Laviada MD, Babín M, Dominguez J, Sánchez-Vizcaíno JM.A sandwich enzyme-linked immunsorbent assay (ELISA) for rapid detection of African horsesickness virus (AHSV) in infected spleens or cell culture supernatant was developed. This method uses two monoclonal antibodies (MAbs) which recognize two non-overlapping epitopes of the major core protein (VP7) to coat the solid phase, and one labeled with biotin as second antibody. This ELISA was evaluated for its ability to detect AHSV in infected spleens resulting in a sensitivity of 97.4% and a specificity of 100% compared with virus isolation in cell culture, and can be used for the detection of the n...
Tsunemitsu H, Jiang B, Saif LJ.Enzyme-linked immunosorbent assays (ELISAs) were developed to detect group (gp) C rotavirus antigens and antibodies. Both assays were confirmed to be specific for gp C rotavirus by using serogroup A, B, and C rotaviruses; hyperimmune antisera to these serogroups of rotaviruses; and paired serum specimens from animals infected with gp C rotaviruses. The ELISA for antigen detection reacted not only with porcine gp C rotaviruses but also with human and bovine gp C rotaviruses. Following experimental challenge of gnotobiotic pigs with porcine gp C rotavirus, the virus was found by ELISA in all dia...
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-...
Hamblin C, Anderson EC, Mellor PS, Graham SD, Mertens PP, Burroughs JN.Four ponies were each inoculated with a different serotype of African horse sickness virus (AHSV) which had been passaged through cell culture in order to achieve attenuation. Three of the ponies died suddenly after showing mild clinical signs, the fourth pony remained clinically normal and was killed at day 38. Infectious AHSV was isolated from blood samples collected at intervals from all four ponies. Positive antigen ELISA reactions were only observed with blood samples from two of the ponies on the two days preceding death. Specific AHSV antibodies were detected by ELISA in serum samples f...
Ball JM, Rushlow KE, Issel CJ, Montelaro RC.We describe here a detailed analysis of the antigenic determinants of the surface unit glycoprotein (gp90) of equine infectious anemia virus (EIAV), using a comprehensive panel of synthetic peptides in enzyme-linked immunosorbent assays with immune serum from naturally and experimentally infected horses and with a panel of gp90-specific neutralizing and nonneutralizing monoclonal antibodies. The results of these studies identify immunoreactive segments throughout the conserved and variable domains of gp90 but localize immunodominant (100% reactivity) determinants to the amino and carboxyl term...
Browning GF, Chalmers RM, Fitzgerald TA, Snodgrass DR.Ten cultivable equine rotavirus isolates, two of North American, six of British, and two of Irish origin, were compared with standard rotavirus strains and with each other by cross neutralization, neutralization with a panel of monoclonal antibodies (MAbs), hybridization to a simian rotavirus (SA-11) VP7 gene probe, and reaction with rotavirus subgrouping and serotyping MAbs in enzyme-linked immunosorbent assays. Six isolates, two of which had previously been serotyped as G3 by other workers, were found to be serotype G3; one was confirmed to be G5, and three were not related to serotypes G1 t...
Lecompte F, Combarnous Y.A simple, accurate, sensitive enzyme immunoassay (EIA) has been developed that permits the measurement of equine Chorionic Gonadotropin activity in pregnant mare plasmas or serums as well as in commercial and highly-purified preparations. This assay is specific for eCG and eLH which share the same polypeptide structure but differ in their oligosaccharidic chains. The more important result is that this EIA has been found to be give data in very close agreement with the in vivo assay. Therefore this very rapid and convenient assay can be used to measure the activity of eCG/PMSG in pregnant mares...
Browning GF, Chalmers RM, Fitzgerald TA, Corley KT, Campbell I, Snodgrass DR.Foal fecal group A rotavirus strains were characterized by electropherotype, serotype, and subgroup and shown to be distinctly different from rotaviruses of other mammals. Of 86 strains that were electropherotyped, 98% had similar profiles, with gene segments 3 and 4 close together and segments 7, 8, and 9 widely spaced. Of 70 strains that had sufficient detectable VP7 antigen to be serotyped by enzyme-linked immunosorbent assays (ELISAs), 63% were serotype G3 (39% were subtype G3A and 24% were subtype G3B), 4% were serotype G13, and 33% were untypeable. Serotypes G1, G2, G4, G5, G6, G9, G10, ...
Mulville P.In the late 1970s, a new infectious disease in horses, involving acute enteritis, was recognised in the Potomac River area of Maryland, U.S.A. The causative agent was identified subsequently as a new species of rickettsial organism, later named Ehrlichia risticii. Since then, the disease has been reported in many other states, and in enzootic areas vaccination is common. Signs associated with the clinical disease included depression, fever, anorexia, decreased or absent intestinal sounds, profuse watery diarrhoea and laminitis. However, considerable variation in clinical manifestations has bee...
Dhawedkar RG, Jain NC, Mount ME, Bowling AT, Vegad JL.An enzyme-linked immunosorbent assay (ELISA) was standardised and applied for the detection of antiplatelet and antineutrophil antibodies using a heterologous system consisting of equine platelets or neutrophils and antisera raised in rabbits. The standardised technique consisted of using Immulon type 3 plate, 1 per cent gelatine as a blocking solution, poly-L-lysine buffer as a coating solution, unfixed antigen, 90 microliters test serum, horseradish peroxidase conjugated antibody and o-phenylenediamine dihydrochloride as a substrate. The number of unfixed platelets or neutrophils required fo...
Stanley S, Jeganathan A, Wood T, Henry P, Turner S, Woods WE, Green M, Tai HH, Watt D, Blake J.We have raised antibodies to morphine and etorphine and developed one-step enzyme-linked immunosorbent assays (ELISA) for these drugs as part of a panel of post race tests for drugs in racing horses. These tests are simple, can be completed in 2 h, and can be read by visual inspection. The morphine ELISA has an I50 for morphine of about 1.5 ng/mL, while the etorphine ELISA has an I50 for etorphine of 250 pg/mL. Cross-reactivity studies show that the antimorphine antibody cross-reacts well with levorphanol, hydromorphone, and oxycodone, while the anti-etorphine antibody showed no cross-reactivi...
Heath SE, Geor RJ, Tabel H, McIntosh K.An enzyme-linked immunosorbent assay (ELISA) was developed for use in horses to determine serum titers of antibodies of the immunoglobulin classes IgA, IgG, and IgM to Streptococcus equi M-like protein and culture supernatant protein antigens. Serum antibodies were determined in 28 adult horses, including 9 horses with recent S. equi infections, 17 horses without known exposure to S. equi, but without a history of respiratory disease in the preceding 4 months, and 2 horses with clinical purpura hemorrhagica. Serum IgA titers to culture supernatant protein antigen were highest in recently infec...
Fitzgerald TA, Browning GF.The sensitivity of a rotavirus serotyping enzyme-linked immunosorbent assay (ELISA) was improved by the addition of 0.5 mM CaCl2 to the washing buffer and reagent diluent. Twenty-nine of 63 (46%) previously untyped bovine and equine faecal rotavirus samples were serotyped in the modified assay. A differential response to Ca2+ ions was noted for different G-serotypes suggesting that serotyping assays performed without the inclusion of CaCl2 in the assay buffers may produce biased results.
Wassall DA, Gregory RJ, Phipps LP.The detection of antibodies against Trypanosoma equiperdum in 689 equid sera was compared by enzyme-linked immunosorbent assay (ELISA), the complement fixation test (CFT) and an indirect immunofluorescent test (IIF). CFT was the least sensitive technique, susceptible to anti-complementary factors and the most technically demanding. IIF was more sensitive, but was only suitable for testing limited numbers of samples. In this study, ELISA was the most sensitive test, the least labour intensive and lends itself to a considerable degree of automation. It is suggested that ELISA would be relatively...
Martens JG, Stephens KA, Kerchner LJ, Heck FC, Martens RJ.Plasma fibronectin concentrations were measured in clinically healthy mares and their neonatal foals, using a modified human fibronectin competitive enzyme-linked immunosorbent assay. Ranges of plasma fibronectin were established in clinically healthy horses, and the assay was reliable and reproducible. Plasma fibronectin concentrations were similar in mares and foals, both before and after colostrum ingestion.
Weiland G, Hasslinger MA, Mezger S, Pöllein W.In an investigation period over 8 months the natural course of infection was studied by means of coproscopic and serological methods in 27 mares and 29 foals. The examination of the stool showed in mares, before the beginning of the grazing season, an infection rate of 100% with small and a rate of 7.4% with large strongyles (Str. vulgaris). Serologically the ELISA showed in foals only a distinct increase of antibody activity with the somatic antigen. The mares retained the high IgG-values of activity, which were already found at the beginning of the investigations. Even though the agglutinati...
Browning GF, Chalmers RM, Sale CS, Fitzgerald TA, Snodgrass DR.The homotypic and heterotypic antibody response to rotavirus was determined in three pony mares and their foals. The normal concentrations of anti-rotavirus antibodies in mares' milk and mares' and foals' serum over the first 10 weeks post-partum were measured using IgA, IgG and rotavirus serotype-specific enzyme linked immunosorbent assays. Experimental infection of the foals with serotype 3 equine rotavirus produced a rapid, serotype-specific response which peaked 10 days after infection and a slower heterotypic response which peaked 32 days later. In contrast, vaccination of the mares with ...
Gaĭdamovich SIa, Pomelova VG, Lavrova NA, Mel'nikova EE, Sokolova MV, Kharitonenkov IG, Zlobin VN.Potentialities of differentiation between Venezuelan equine encephalomyelitis (VEE) complex viruses by time-resolved fluoroimmunoassay and enzyme immunoassay were studied. For this, 4 test systems were used based on different combinations of native and labeled polyclonal antibodies to VEE virus, strain Trinidad, and monoclonal (MCA) antibody MAK 14-7 to protein EL of this virus. The maximal sensitivity and specificity was achieved in the test system formed from native MCA MAK 14-7 for sensitization of the solid phase and labeled polyclonal immunoglobulins for demonstration of the test results....
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...
Kirkpatrick JF, Shldeler SE, Lasley BL, Turner JW.This study was carried out to develop an accurate, rapid and inexpensive method for diagnosing pregnancy in uncaptured feral horses by analysis of fecal steroid metabolites and to compare the accuracy of this method with diagnosis by urinary estrone conjugates (E(1)C). Paired urine and fecal samples were collected from 40 sexually mature feral mares during August and October. Urine samples were extracted directly from the soil and analyzed by enzymeimmunoassay (EIA) for E(1)C. Water extracts of fecal samples were assayed by EIA for E(1)C and nonspecific progesterone metabolites (iPdG). Urinary...
Payne RC, Sukanto IP, Djauhari D, Partoutomo S, Wilson AJ, Jones TW, Boid R, Luckins AG.Cattle, buffaloes and horses in several areas of Indonesia were examined for evidence of infection with Trypanosoma evansi by the microhaematocrit centrifugation technique (MHCT) and an enzyme-linked immunosorbent assay (ELISA) for detection of antibodies to T. evansi. Evidence of infection was found in animals at each sampling site although differences were seen in prevalence rates between sites. Prevalence rates in buffalo were usually higher than in cattle in the same area while in horses they were much lower than in cattle or buffalo. An age-dependent prevalence rate was seen in buffalo an...
Chong YH, Ball JM, Issel CJ, Montelaro RC, Rushlow KE.Defined segments of the transmembrane envelope glycoprotein (gp45) of equine infectious anemia virus were expressed as TrpLE fusion proteins and examined for their reactivity in Western immunoblots against a diverse panel of equine immune sera. The most immunogenic region of gp45 was localized to its amino terminus, positioned between the hydrophobic fusion and the transmembrane domains. A series of overlapping synthetic peptides were used in enzyme-linked immunosorbent assays to define an immunodominant epitope within this region. In contrast, the carboxy-terminal half of gp45 displayed both ...
Berti A, Tremori E, Pazzagli L, Degl'Innocenti D, Camici G, Cappugi G, Manao G, Ramponi G.Acylphosphatase was purified from rat skeletal muscle essentially by gel filtration and high-performance ion-exchange chromatography. The complete amino acid sequence was reconstructed by using the sequence data obtained from tryptic, peptic, and S. aureus V8 protease peptides. The protein consists of 96 amino acid residues and is acetylated at the NH2-terminus. The immunological cross-reactivity of acylphosphatase from rat and horse skeletal muscle was examined by ELISA. The reaction with rabbit antiserum revealed the presence of at least five antigenic sites on rat enzyme, two of which are c...
Mazzei M, Savini G, Di Gennaro A, Macchioni F, Prati MC, Guzmàn LR, Tolari F.West Nile virus (WNV) is a mosquito-borne virus belonging to the family Flaviviridae included in the Japanese encephalitis antigenic complex (JEAC). A seroepidemiological study was carried out in 2011 using 160 horse sera collected from different areas of Bolivia to investigate the presence of WNV antibody. A high proportion (59.4%) of the tested sera were positive to a commercially available WNV competitive enzyme-linked immunosorbent assay (C-ELISA). Sixty-six randomly selected C-ELISA-positive sera were further tested by WNV plaque reduction neutralization test (PRNT), virus neutralization ...
Hannan MA, Murata K, Takeuchi S, Haneda S, Cheong SH, Nambo Y.The objectives of this study were to determine the plasma profile of equine chorionic gonadotropin (eCG) and its association with the formation of supplementary corpus luteum (CL) and plasma progesterone concentrations in embryo transfer Hokkaido native pony recipient mares. Blood samples and transrectal ultrasound examination of the reproductive tract were carried out weekly from the day of ovulation until week 32 of gestation (n = 4). Plasma concentrations of eCG and progesterone were measured by enzyme immunoassays. The eCG concentration was first detectable at week 5 for 2 mares and at wee...
Cavalcanti RTC, Teixeira PAC, Levy RS, Pereira HMG, Aquino Neto FR.Erythropoiesis-stimulating agents (ESAs) have been used in horses for doping purposes to increase the performance of these animals in endurance sports. Currently, enzyme-linked immunosorbent assay (ELISA) and mass spectrometry methods are used to detect ESA abuse in equines. However, the sarcosyl polyacrylamide gel-electrophoresis (SAR-PAGE) technique could also be used, since its application in human doping control is well established and has proven to be more sensitive. In this work, the SAR-PAGE method was used to detect recombinant human erythropoietin (rHuEPO), novel erythropoiesis stimul...
Chung C, Wilson C, Timoney P, Adams E, Adams DS, Chung JS, Evermann JF, Shuck K, Lee SS, McGuire TC.Equine arteritis virus (EAV) causes contagious equine viral arteritis, characterized by fever, anorexia, conjunctivitis, nasal discharge, dependent edema, abortion, infrequent death in foals, and establishment of the carrier state in stallions. The World Organization for Animal Health (OIE) defines a horse as seropositive if the serum neutralization (SN) antibody titer is ≥1:4 to EAV. However, determining the SN titer is time-consuming and requires specific laboratory facilities, equipment, and technical expertise to perform. Furthermore, interpretation of the SN titer of some sera can be di...
Mariella J, Castagnetti C, Prosperi A, Scagliarini A, Peli A.The purpose of this study is to investigate the presence of IL-4, IL-8, IL-13 and IFN-γ in equine colostrum and in foals' serum. Samples were obtained from 14 mares and their healthy foals. Soon after parturition, 10ml of colostrum was collected, filtered, centrifuged and frozen until assayed. Blood samples were obtained from each foal at birth (TO) and again after 24h (T24), after which they were frozen until assayed. Serum IgG was measured at 24h of age with an immunoturbidimetric quantitative method. Cytokine concentration was determined using commercially available ELISA tests. Statistica...
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 ...
Uzcanga GL, Bubis J.Two horses were infected with distinct non-tsetse transmitted Venezuelan stocks, namely TeAp-N/D1 and TeAp-El Frio01 . Preceding reports have revealed that a 64-kDa antigenic glycopolypeptide (p64), which is the soluble form of the predominant variant surface glycoprotein from TeAp-N/D1 , can be used as a good antigen for immunodiagnosis of animal trypanosomosis. Here, the course of the experimental acute infection in both horses was monitored by evaluating total anti-p64 IgG and particular anti-p64 γ-specific IgG and μ-specific IgM isotypes in sera using indirect enzyme-linked immunoso...
Rosskopf U, Noeske K, Werner E.This paper describes a double antigen ELISA (DAE) for rapid, specific and reliable assessment of the antitetanus immune status of horses and sheep. Compared with the indirect ELISA, the double antigen ELISA has the advantage of species-independent testing of sera. Thanks to its test design, it is more specific since the detected antibodies are forced to bind tetanus toxoid twice. In addition, it is very sensitive to tetanus antibodies, enabling the detection of low antibody titres, in range which is relevant for the assessment of the protective status (tetanus toxin neutralising antibodies). T...
Thougaard AV, Jaliashvili I, Christiansen M.The glycoprotein tetranectin (TN) found in human serum is a 90-kDa homotrimeric C-type lectin binding Ca2+, heparin and plasminogen kringle 4. TN is suggested as being implicated in tissue remodelling. The antigenic reactivity of putative TN was examined in serum from 14 different animal species using three sandwich enzyme immunoassays for human TN. Crab-eating macaque serum showed the strongest reaction, followed by horse and cat. Serum from cow, goat, pig, mouse and chicken reacted weakly, while dog, trout, and the amphibian and the reptile species did not react. The TN-like protein from mac...
Mar Htun Z, Laikul A, Pathomsakulwong W, Yurayart C, Lohnoo T, Yingyong W, Kumsang Y, Payattikul P, Sae-Chew P, Rujirawat T, Jaturapaktrarak C....Pythium insidiosum causes a life-threatening infection termed pythiosis in humans and other animals. The organism has been identified in tropical and subtropical environments worldwide. Since 1985, human pythiosis has been increasingly reported from Thailand. Seroprevalence studies estimated that 32,000 Thai people had been exposed to the pathogen. In 2018, the first animal pythiosis case in Thailand was diagnosed in a horse. Here, we investigated the seroprevalence of anti-P. insidiosum antibodies in the Thai equine population. Methods: We surveyed serum anti-P. insidiosum antibodies in 15...
Suter M, Fey H.An enzyme-linked immuno sorbent assay (ELISA) for measuring horse IgE specific to ovalbumin, bencylpenicilloic acid and odinitrocarboxyphenol is described. We used a sandwich type of ELISA by which horse serum was incubated in antigen-coated tubes containing one additional polystyrene ball, followed by rabbit anti horse IgE serum. The tubes were then incubated with biotinylated goat anti rabbit globulin followed by avidin coupled to phosphatase. Endpoint titrations were compared. The ELISA is highly reproducible due to the pretreatment of the polystyrene with glutaraldehyde. The increased anti...
Hu WG, Chau D, Wong C, Masri SA, Fulton RE, Nagata LP.The genes encoding envelope proteins E1 and E2 of western equine encephalitis virus (WEEV) were respectively cloned into a prokaryotic T7 RNA polymerase-regulated expression vector. The recombinant C-terminal 6xHis-tagged WEEV E1 and E2 were expressed in bacteria as inclusion bodies that were subsequently solubilized with 8M urea, purified by immobilized metal ion affinity chromatography and finally refolded using an arginine system. The purified 6xHis-tagged proteins showed 50kDa bands as revealed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis, consistent with the expected size...
Chang Y, Maylin GM, Matsumoto G, Neades SM, Catlin DH.Methods have been developed to screen for and confirm darbepoetin alfa, recombinant human EPO, and methoxy polyethylene glycol-epoetin β (PEG-epoetin β) in horse plasma. All three methods screen samples with an enzyme-linked immunosorbent assay (ELISA) and confirm by liquid chromatography-tandem mass spectrometry (LC-MS/MS). This report focuses on PEG-epoetin β. The ELISA assay was able to detect PEG-epoetin β at 0.02 ng/mL in 50 µL of horse plasma. Many samples had high background levels of immunoreactivity; however, introducing polyethylene glycol 6000 (PEG 6000) into the samples before...
Guillot J, Sarfati J, Ribot X, Jensen HE, Latgé JP.To detect antibodies against Aspergillus fumigatus antigens in serum samples from horses and to evaluate the relevance of this method as an alternative approach to the diagnosis of mycosis of the auditory tube diverticulum (guttural pouch mycosis [GPM]). Methods: Twelve clinically normal horses (controls) and 12 horses with GPM diagnosed by endoscopic observation of characteristic mycotic plaques. Methods: Antibodies to A fumigatus antigens were detected in serum by use of an ELISA and immunoblot analysis with extracellular antigens. Results: Antibodies against A fumigatus antigens were found ...
Gilkerson JR, Love DN, Drummer HE, Studdert MJ, Whalley JM.To investigate the seroprevalence of equine herpesvirus 1 in foals around weaning and after weaning on two large Thoroughbred farms using a type-specific enzyme-linked immunosorbent assay to determine exposure to infection. Methods: A longitudinal population study in groups of Thoroughbred weanling foals. Methods: Two hundred weanling Thoroughbred foals from a population of about 380 foals were enrolled on two adjacent stud farms in the Hunter Valley of New South Wales. Foals on both farms were weaned from February to May 1995 into randomly selected groups of 10 to 15 foals. Farms were selecte...
Berti A, Tremori E, Pazzagli L, Degl'Innocenti D, Camici G, Cappugi G, Manao G, Ramponi G.Acylphosphatase was purified from rat skeletal muscle essentially by gel filtration and high-performance ion-exchange chromatography. The complete amino acid sequence was reconstructed by using the sequence data obtained from tryptic, peptic, and S. aureus V8 protease peptides. The protein consists of 96 amino acid residues and is acetylated at the NH2-terminus. The immunological cross-reactivity of acylphosphatase from rat and horse skeletal muscle was examined by ELISA. The reaction with rabbit antiserum revealed the presence of at least five antigenic sites on rat enzyme, two of which are c...
Hopke K, Coleman M, Kneese E, Dominguez B, Diesel A, Patterson A.The existence of antibodies against cross-reactive carbohydrate determinants (CCDs) has been studied extensively in humans, and more recently, in dogs and cats. These antibodies can reduce the specificity of in vitro serum allergen tests. Objective: To investigate the prevalence of anti-CCD immunoglobulin (Ig)E in both allergic and nonallergic horses as well as evaluate its potential impact on serum allergen testing. Methods: Twenty-one allergic and 21 nonallergic horses. Methods: Sera were analysed for anti-CCD IgE utilising a commercial CHO enzyme-linked immunosorbent assay (ELISA)...
McCarthy MB, Duesterdieck-Zellmer KF, Larson MK.To determine whether Botulinum neurotoxin type A (BoNT-A) ameliorates the effects of interleukin 1 (IL-1) on equine articular cartilage, or exerts negative effects on normal equine articular cartilage homeostasis in vitro. Methods: Articular cartilage explants from 6 healthy femoropatellar joints of 3 adult horses. Methods: Explants were allocated to the IL-1 challenged or unchallenged group, then exposed to 1 of 6 concentrations of BoNT-A (0, 1, 10, 50, 100, or 500 pg/mL) for 96 hours. To assess BoNT-A's effects on inflammation, prostaglandin E2 (PGE2) was measured in media via ELISA. Matrix ...
Borst GH, Smidt WJ, Berghuis GA.The concentrations of progesterone in milk were determined in twenty-one mares to establish a diagnosis of pregnancy in an early stage (15-19 days). Progesterone levels varied from 0.0 to 4.2 ng/ml in nine non-pregnant mares and from 6.7 to 30.0 ng/ml in twelve pregnant mares. Progesterone levels were determined by an enzyme-linked immunosorbent assay (ELISA).
Hemida MG, Alhammadi M, Daleb A, Alnaeem A.African horse sickness virus (AHSV) is one of the most devastating viral diseases of the family Equidae. Infection with AHSV threatens not only the Saudi equine industry but also the equine industry worldwide. This is due to the high morbidity and mortality rates among the infected population of up to 100%. The World Organisation for Animal Health (OIE) lists AHSV among its notifiable diseases; this requires Member Countries to monitor the situation with regard to AHSV very carefully in order to avoid the spread of the virus. The OIE also suggests the systematic monitoring of AHSV in the equin...
Zahradnik E, Sander I, Lotz A, Liebers V, Thullner I, Tacke S, Raulf M.The study aimed to determine the allergen, endotoxin and β-(1,3)-glucan concentrations at various areas on a university campus of veterinary medicine. Methods: Dust samples were collected four times a year for three years using electrostatic dust collectors (EDC) at 25 different locations on a campus of veterinary medicine and in laboratories of inorganic chemistry as a control area representing animal-free environment. Major animal allergens from dog, cat, horse, cattle and mouse, domestic mite (DM) allergens, and β-(1,3)-glucan were measured using enzyme immunoassays and endotoxin using th...
Schusser GF, Köller G, Recknagel S, Lindner A.The hay producing plants, concentrate, straw and meadows could be contaminated by the aerosols of glyphosate based herbicide during spraying process of crops and pre-harvest desiccation treatment of cereals. The aim of this study is to investigate the concentration of glyphosate in the duodenal fluid of horses with gastric ulcer syndrome. The stomach and duodenum of referred untreated horse patients (n=92) with colic, weight loss, diarrhoea, anemia or performance intolerance were endoscopically examined right after the admission. Duodenal fluid (40 ml) was collected from the duodenal region ...
Nishita T, Goto T, Kimura H, Asari M.1. In this study, carbonic anhydrase III (CA-III) content in 18 equine muscles was determined by enzyme immunoassay. 2. It was found to differ in several muscles. 3. That in external intercostal muscle, rectus abdominis muscle and splenius muscle from four horses was very high. 4. Although the masseter muscle had only type I fibers, CA-III content was similar to that in mixed-fiber type muscles such as the biceps femoris muscle. 5. It thus appear that equine type I fibers can be further subgrouped.
Andraus MH, Siqueira ME.An analytical procedure to screen butorphanol in horse race urine using ELISA kits and its confirmation by GC-MS is described. Urine samples (5 ml) were subjected to enzymatic hydrolysis and extracted by solid-phase extraction. The residues were then evaporated, derivatized and injected into the GC-MS system. The ELISA test (20 microl of sample) was able to detect butorphanol up to 104 h after the intramuscular administration of 8 mg of Torbugesic, and the GC-MS method detected the drug up to 24 h in FULL SCAN or 31 h in the SIM mode. Validation of the GC-MS method in the SIM mode using nalbup...
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 ...
Harkiss GD.An isotype-specific microELISA is presented for the measurement of antibodies to equine antithymocyte globulin in human heart transplant recipients. The assay conditions were optimized and evaluated in serial samples from 40 patients receiving a cardiac allograft. The results demonstrate that despite steroid immunosuppression and T cell cytopenia the majority of patients receiving antithymocyte globulin develop significant antibody responses, with some producing very high titres. IgM and IgG isotypes tended to predominate, with peak antibody responses occurring during the second and third week...
Lettry V, Sumie Y, Mitsuda K, Tagami M, Hosoya K, Takagi S, Okumura M.The objective of this study was to investigate the changes in synovial fluid concentration of collagen type II cleavage site (C2C) and procollagen II C-propeptide (CPII), markers of joint cartilage degeneration and synthesis, respectively, in horses with intraarticular fracture or osteochondrosis dissecans (OCD), and to examine the relationship between arthroscopic findings and these biomarker levels. Synovial fluid was collected from 36 joints in 18 horses (6 fractures and 12 OCDs). Samples from contralateral normal joints, when available, served as controls (n = 12). Concentrations of C2C an...
