Laboratory methods in equine research encompass a variety of techniques and procedures used to analyze biological samples from horses to study health, disease, genetics, and physiology. These methods include hematological analyses, biochemical assays, molecular biology techniques, and microbiological cultures. Commonly utilized laboratory techniques involve blood tests for complete blood count (CBC) and serum chemistry, polymerase chain reaction (PCR) for genetic and infectious disease studies, and enzyme-linked immunosorbent assays (ELISA) for detecting specific proteins or antibodies. These methods provide valuable data that contribute to understanding equine health and disease mechanisms. This page compiles peer-reviewed research studies and scholarly articles that explore the application, development, and outcomes of laboratory methods in the context of equine research.
McDowell KJ, Adams MH, Williams NM.Proteins synthesized and released in vitro by oviducts collected from horse mares during estrus and at day 4 after ovulation for bred and nonbred mares were examined by two-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis (2-D SDS PAGE) and fluorography. Ampullary and isthmic regions both produced a wide array of nondialyzable proteins in culture. Major proteins or groups of proteins identified according to relative molecular weight (kDa) and apparent isoelectric point (pI) were at 100 kDa, pI 8; 100-200 kDa, pI 6; 150 kDa, pI 4.5; 60-100 kDa, pI 4; and an array of polypep...
Whitehair KJ, Steffey EP, Willits NH, Woliner MJ.To study behavioral and cardiopulmonary characteristics of horses recovering from inhalation anesthesia, 6 nonmedicated horses were anesthetized under laboratory conditions on 3 different days, with either halothane or isoflurane in O2. Anesthesia was maintained at constant dose (1.5 times the minimum alveolar concentration [MAC]) of halothane in O2 for 1 hour (H1), halothane in O2 for 3 hours (H3), or isoflurane in O2 for 3 hours (I3). The order of exposure was set up as a pair of Latin squares to account for horse and trial effects. Circulatory (arterial blood pressure and heart rate) and re...
Fjeldsgaard BE, Paulsen BS.Extracts of horse dander (HD) and horse hair and skin scrapings (HHSS) have been compared with respect to their content of proteins and carbohydrates. The protein content of HD is more than double that of HHSS, while the carbohydrate content is of the same order. SDS-PAGE and IEF, both combined with immunoblotting, and CIE/CRIE showed the IgE-binding ability of the proteins/glycoproteins present in the two extracts. SDS-PAGE/immunoblotting showed the presence of mainly the same IgE-binding bands in the two extracts. Nine were detected in HD, and seven in HHSS. Four of these were glycoproteins....
Wilson SM, Pediani JD, Ko WH, Bovell DL, Kitson S, Montgomery I, Brown UM, Smith GL, Elder HY, Jenkinson DM.When sweat glands isolated from samples of horse skin were explanted and cultured under favourable conditions, they could exhibit cellular outgrowth. This growth could be maintained for 2-4 weeks and these primary cultures were then disaggregated and the resultant cell suspensions used to initiate epithelial cell lines. Secretion from intact equine sweat glands is regulated by beta 2-adrenoceptors and appears to be mediated by cyclic AMP, but there is evidence that calcium may also play a role. Adrenaline could increase the cyclic AMP content of the cultured cells and this response was mediate...
Auer DE, Skelton KV, Tay S, Baldock FC.Total plasma carbon dioxide (TCO2) concentrations were measured in Standardbred horses to determine criteria to discriminate between normal horses and horses with excessive TCO2 concentrations on raceday. TCO2 concentrations from stabled horses were distributed normally with a mean of 30.2 mmol/L and a standard deviation of 1.2 (n = 192) while pre-race TCO2 concentrations were not normally distributed. The results indicate that about 50 horses per million are likely to have TCO2 concentrations greater than or equal to 35 mmol/L and that it is extremely unlikely that a normal horse would have a...
Boschwitz JS, Timoney JF.A rapid method for purifying equine C3 which yields milligram quantities of pure C3 is described. Protein from equine plasma was selectively precipitated with polyethylene glycol, and the C3 was purified by anionic and cationic exchange HPLC. The yield from this procedure was 12%. The purified C3 was composed of an alpha chain (118 kD) and a beta chain (68 kD) linked by at least one disulfide bond, and it had an isoelectric point of 4.7. Amino acid analysis indicated a strong conservation of amino acid usage between equine and human C3. The N-terminal sequences of the alpha and beta chains wer...
Richa , Grover YP, Charan S.The authors describe a rapid and simple dot immunobinding assay (DIA) for detection and identification of equine herpesvirus-1 antigen in field samples from cases of abortion, stillbirth, perinatal foal mortality and paralysis. The assay employs a nitrocellulose membrane to which antigen is adsorbed as a dot. Antigen is identified as a coloured dot using a procedure based on the principle of enzyme-linked immunosorbent assay (ELISA). In all, 61 samples were tested by DIA and the test was compared with conventional agar gel immunodiffusion (AGID). With DIA, 44 (72%) samples gave positive result...
Ellington JE, Ignotz GG, Varner DD, Marcucio RS, Mathison P, Ball BA.Coculture of stallion sperm with monolayers of equine oviductal epithelial cells (OEC) was evaluated. Monolayers were obtained from frozen-thawed OEC. Live sperm attached to the OEC in vitro, whereas sperm killed by heat treatment or glutaraldehyde fixation did not. Sperm attached to OEC showed flagellar motion for 4 d in vitro, during which time they gradually became released. Scanning electron-micrographs showed an intimate association between the sperm and OEC. Incubation of sperm for 4 h with either control, heparinized or OEC-conditioned medium (Tyrode's albumin lactate phosphate) resulte...
Komori M, Higami A, Imai Y, Imaoka S, Funae Y.A form of P450 [termed P450(h-1)] was purified from the liver microsomes of a male horse to electrophoretic homogeneity. The specific content of the final P450(h-1) preparation was 14.8 nmol/mg of protein and the recovery was 0.38% of the microsomal P450. The apparent molecular weight of P450(h-1) was 52,000 Da. The absorption spectra of P450(h-1) indicated that P450(h-1) was a low- and high-spin mixed type P450 in the oxidized form. The reconstituted system containing P450(h-1) could catalyze benzphetamine N-demethylation, 7-ethoxycoumarin O-deethylation, and testosterone 16 alpha-hydroxylati...
Messick JB, Rikihisa Y.The binding, internalization, and proliferation of Ehrlichia risticii in P388D1 cells and equine polymorphonuclear (PMN) leukocytes were studied by immunofluorescent staining and flow cytometric analysis. The binding of ehrlichiae to P388D1 cells at 4 degrees C was dose dependent, and the antigens of bound organisms were susceptible to pronase treatment. Additionally, the binding of ehrlichiae to P388D1 cells was diminished when either P388D1 cells or ehrlichiae were treated with 1% paraformaldehyde for 30 min or 0.25% trypsin for 15 min. These results indicate that the ehrlichial ligand and h...
Takeda S, Ohta M, Ebina S, Nagayama K.Horse L-ferritin cDNA was cloned from horse liver, and the base sequence was determined. The L-ferritin was expressed using pTZ18U encoding lac promoter, and found to possess an additional 8-amino acid sequence at the N-terminus as compared with commercially obtained horse spleen (natural) ferritin. It was determined that there was Pro at position 94 in both the recombinant and natural L-ferritin, although it was previously reported that Leu was in this position in the natural species. Transmission electron microscopy showed that this recombinant ferritin formed a 24-mer shell.
Bolann BJ, Ulvik RJ.The redox interaction between O2.- and ferritin cannot solely be regarded as as a Fe(II) release reaction. We demonstrate that native copper bound to horse spleen ferritin and apoferritin, stimulated the decay of O2.- in a catalytic reaction. Copper was determined by atomic absorption spectrophotometry. Decay of O2.- was monitored spectrophotometrically as the decrease in (A250-A360) at pH 9.5. The catalytic effect was linearly related to the copper content of the protein. Ferritin copper was less efficient than equimolar CuCl2, and iron-poor ferritin was more efficient than iron-rich ferritin...
Strange R, Morante S, Stefanini S, Chiancone E, Desideri A.Extended X-ray absorbance fine structure measurements have been carried out on the initial Fe(III)-apoferritin complex at a Fe/subunit ratio of 2 in native and modified horse spleen apoferritin. Analysis of the data indicates that in the native protein the iron forms a protein-bound polynuclear cluster (Fe-Fe distance 3.4 A) with a first coordination sphere constituted by 5-6 low-Z atoms, e.g., nitrogen atoms, carboxylate-like ligands or oxo bridges between the iron atoms. Modification of Cys-126, a residue localized on the outer surface of the hydrophilic three-fold channels, with p-chloromer...
Menzeleev RF, Smirnova GP, Chekareva NV, Zvonkova EN, Krasnopol'skiĭ IuM, Shvets VI.An increase of the mouse fibroblast proliferation by ganglioside GM3 from equine erythrocytes is described. The structure of GM3 has been established on the basis of chemical methods, enzymatic degradation, GC-MS, as well as plasma desorption mass spectrometry and HPLC of 9-anthrylmethyl esters of gangliosides to characterize the long-chain base composition. The oligosaccharide moiety includes an N-glycolylneuraminic acid residue, whereas the main components of the lipid moiety are 20:1 sphingosine and 24:0 fatty acids.
Gregg AS, Jones RS, Snowdon SL.A fault in the assembly of a Matrix Large Animal Circle anaesthetic machine resulted in reversal of fresh gas flow through the vaporizer. The fault was discovered only after the sudden development of excessive depth of anaesthesia in two equine patients. Laboratory investigations were conducted to determine the effect of flow reversal on vaporizer output. Results indicated that output concentration was approximately doubled under these conditions.
Santucci R, Ascoli F, La Mar GN, Pandey RK, Smith KM.The reconstitution kinetics of horse heart myoglobin, as met-cyano derivative, with two synthetic hemins in which the 6- or the 7-propionate is replaced by a methyl group, has been investigated by circular dichroism, in order to gain information on the heme re-orientation process following the heme insertion into the globin pocket. The results obtained confirm that the preferred heme orientation places the sole propionate into the position occupied by the 6-propionate in the crystal structure, supporting the importance of the salt bridge occurring between this propionate and the basic CD3 resi...
Casey PJ, Hillman RB, Robertson KR, Yudin AI, Liu IK, Drobnis EZ.An acrosomal staining technique that can differentiate between living and dead sperm was developed for equine sperm. The fluoresceinated lectin Pisum sativum agglutinin (FITC-PSA) was used to identify the presence or absence of acrosomal contents, while the supravital nuclear dye Hoechst 33258 (H258) was used to assess viability. The accuracy of the FITC-PSA acrosomal stain was tested by comparing the percentage of sperm that had lost their acrosomal contents, detected by the staining method, with that detected by transmission electron microscopy (TEM). Following capacitation in vitro, the acr...
McGorum BC, Dixon PM, Halliwell RE, Irving P.The urea and albumen dilution techniques for standardising the variable concentrations of pulmonary epithelial lining fluid (PELF) in bronchoalveolar lavage fluid (BALF) samples were evaluated in horses. Both techniques proved satisfactory and were of equal accuracy. Albumen adjusted BALF cell counts were significantly higher than PELF cell counts. BALF from normal horses contained, on average, 0.4 per cent PELF (range 0.1 to 1.0 per cent), as determined by the urea dilution technique.
Harty RN, Caughman GB, Holden VR, O'Callaghan DJ.Equine herpesvirus 1 (EHV-1, Kentucky A strain) preparations enriched for defective interfering particles (DIPs) can readily establish persistent infection. The UL1 gene, which is conserved in the genome of DIPs that mediate persistent infection, maps between nucleotides 1418 and 2192 (258 amino acids) from the L (long) terminus. UL1 has no homology with any known gene encoded by herpes simplex virus type 1 but has limited homology to open reading frame 2 of varicella-zoster virus and the "circ" gene of bovine herpesvirus type 1. Previous work showed that the EHV-1 UL1 gene belongs to the earl...
Dennis VA, Klei TR, Chapman MR.Supernatants generated by stimulation of peripheral blood mononuclear cells (PBMC) from Strongylus vulgaris sensitized or immunized ponies were assayed in vitro for eosinophil chemotactic activity (ECA) using the filter system in blind well chambers. The supernatants from these cultures were chemotactic for eosinophils, but not for neutrophils. Supernates from cultures of unsensitized PBMC stimulated with S. vulgaris antigen were not chemotactic for eosinophils. ECA was first detected in culture supernatants after 1.5 h of incubation and was dependent on both antigen and PBMC concentrations, b...
Delbeke FT, Teale P, Debackere M, Houghton E.A commercially available generic promazine ELISA kit is available which shows cross-reactivity for the tranquilizer chlorprothixene (CPT). The ELISA test readily detects the presence of CPT or its metabolites in equine urine for up to 24 h after the i.v. and i.m. administration of sub-therapeutic doses (4.5 mg) to three horses. Maximum concentrations (CPT equivalents) are obtained 2 h after i.v. dosing. No distinct concentration peak values are observed after i.m. administration. Following solid-phase extraction, confirmation of CPT and its metabolites by electron impact mass spectrometry afte...
Stockert JC, Trigoso CI, Tato A, Ferrer JM.The structure of specific granules from horse eosinophil leukocytes is still largely unknown. In this work, electron microscopical studies of horse eosinophils reveal that the large cytoplasmic granules contain an external membrane, a matrix of less density, and a dense (non crystalline) core. Round vacuolar inclusions of matrix materials were often observed within the cores. Horse eosinophil granules showed a considerable heterogeneity, and three morphological types could be identified according to structural features of the core and matrix.
Ho JX, Holowachuk EW, Norton EJ, Twigg PD, Carter DC.The amino-acid sequence and three-dimensional structure of equine serum albumin have been determined. The amino-acid sequence was deduced from cDNA isolated from equine liver. Comparisons of the primary structure of equine serum albumin with human serum albumin and bovine serum albumin reveal 76.1% and 73.9% sequence identity, respectively. The three-dimensional structure was determined crystallographically by the molecular-replacement method using molecular coordinates from the previously determined structure of human serum albumin, to a resolution of 0.27 nm. In accordance with the primary s...
Rikihisa Y, Wada R, Reed SM, Yamamoto S.The role of the humoral immune response in ehrlichial infection is unknown. Development of neutralizing antibodies during a course of Ehrlichia risticii infection in a pony was examined in vitro by determining the inhibition of E. risticii infection of P388D1 cells in the presence of the sera. The pony experimentally infected with E. risticii developed significant neutralizing activity in the sera by 15 days postinfection when parasitemia started to decline. Neutralizing activity continued to rise after recovery from the disease up to 34 days postinfection at which time the experiment was term...
Scudamore CL, Pemberton A, Watson ED, Miller HR.Studies have demonstrated that as a result of proteolytic inactivation or complex formation (with neutrophil elastase), human alpha-1-proteinase inhibitor (API) becomes a potent chemoattractant for human neutrophils. The present study aimed to investigate the in vitro chemotactic response of equine neutrophils to an equivalent complex of equine API and neutrophil elastase. No evidence of neutrophil migration was observed towards purified complex derived from equine neutrophil elastase and the Spi 1 isoform of equine API, or to crude mixtures of porcine pancreatic elastase and unseparated equin...
Caron JP, Toppin DS, Block JA.Middle carpal cartilage explants from 4 horses with mild osteoarthritis involving that joint were maintained in tissue culture to test the effects of a polysulfated glycosaminoglycan (PSGAG) on proteoglycan synthesis and degradation. Cultures were exposed to 0.025 or 25 mg of PSGAG/ml for 48 hours, after which the medium was replaced with medium containing similar doses of PSGAG and 35S. Subsequently, the sulfated proteoglycan content of the medium and extracts of the explants was measured. Gel filtration chromatography was used to estimate the size and to purify the principal, large proteogly...
Afshar A, Shakarchi NH, Dulac GC.Two competitive (C) enzyme-linked immunosorbent assays (ELISAs) were developed for the detection of antibodies to vesicular stomatitis virus (VSV) in animal sera. The assays are based upon the availability of polyclonal antibodies (PAbs) from mouse ascitic fluids prepared against the New Jersey (NJ) and the Indiana (IN) VSV serotypes. The assays were performed by the immobilization of VSV-NJ and VSV-IN antigens on a solid phase (microtiter plate). Appropriately diluted test serum mixed with an equal volume of serotype-specific PAb was allowed to incubate in the presence of the relevant VSV ant...
Hope E, Johnston SD, Hegstad RL, Geor RJ, Murphy MJ.A commercially available radioimmunoassay kit for measurement of human osteocalcin was validated for use in horses. For accurate measurement of equine serum osteocalcin, blood samples may be collected at a temperature between 20 and 25 C, then centrifuged within 90 minutes; serum may be stored at -20 C in plastic tubes for up to 26 weeks. Serum may be thawed and refrozen up to 5 times without significant change in measured equine serum osteocalcin concentration. Assay sensitivity was 0.16 ng/ml. Recovery of bovine osteocalcin standard added to equine serum was linear. Intra-assay coefficient o...
Nett TM, Holtan DW, Estergreen VL.The levels of LH, prolactin and oestrogen were measured by radioimmunoassay in serum obtained from six mares during the post-partum period, and LH was also assayed in blood samples collected at 6-hr intervals from four normal cyclic mares during oestrus and at daily intervals during dioestrus. The concentrations of oestrogen and LH suggested that the mares were having ovulatory cycles during the post-partum period irrespective of behavioural oestrus, and the levels of prolactin seemed to rise in association with probable ovulatory LH peaks. In these mares levels of LH had increased significant...
Stanley SM, Kent S, Rodgers JP.In this study the equine metabolism of fluoxymesterone (9alpha-fluoro-11beta-17beta-dihydroxy-17alpha-meth ylandrost-4-ene-3-one) given orally has been investigated. The parent material was not detected, but two major 16-hydroxy metabolites which corresponded to a mono- and a di-hydroxylation product were evident. One of the hydroxylation positions was identified as C-16. Phase II metabolism in the form of glucuronide formation was also common. These steroids will provide target compounds for confirming abuse of this drug in the horse.
Xijier , Mori Y, Fukuoka M, Cairangzhuoma , Inagaki M, Iwamoto S, Yabe T, Kanamaru Y.Native alpha-lactalbumins (α-LA) from equine, bovine, and human milk were not cytotoxic. However, after treatment with trifluoroethanol (TFE), all three α-LAs exhibited cytotoxicity. Toxic potencies were distinctly different among them. Equine α-LA was the most robust, bovine α-LA was moderate, and human α-LA was weak. There were no significant structural changes as between the native and the TFE-treated α-LAs.
How SJ, Lloyd DH, Lida J.A monoclonal antibody (McAb) to Dermatophilus congolensis was produced from murine hybridoma cultures and purified by affinity chromatography. Species specificity was demonstrated using indirect immunofluorescent staining; the McAb was shown to react with 10 D congolensis isolates but not with 10 Nocardia species isolates, a Rhodococcus and a Streptomyces species isolate. The McAb was used to demonstrate D congolensis in clinical material from confirmed bovine and ovine cases and presumptive equine cases of dermatophilosis by indirect immunofluorescent staining.
Lehner AF, Hughes CG, Karpiesiuk W, Harkins JD, Dirikolu L, Bosken J, Camargo F, Boyles J, Troppmann A, Woods WE, Tobin T.Amitraz (N'-(2,4-dimethylphenyl)-N-[[(2,4-dimethylphenyl)imino]methyl]-N-methyl-methanimidamide) is an alpha-2 adrenergic agonist used in veterinary medicine primarily as a scabicide- or acaricide-type insecticide. As an alpha-2 adrenergic agonist, it also has sedative/tranquilizing properties and is, therefore, listed as an Association of Racing Commissioners International Class 3 Foreign Substance, indicating its potential to influence the outcome of horse races. We identified the principal equine metabolite of amitraz as N-2,4-dimethylphenyl-N'-methylformamidine by electrospray ionization(+...
Malmheden Yman I, Sandberg K.Meat from the species horse, donkey and their hybrids, mule/hinny, can be reliably identified by determination of genetic variants of serum albumin by starch gel electrophoresis of meat extracts. Staining of the starch gel for carboxylesterase activity permits differentiation of most horses from donkeys while mules/hinnies cannot be distinguished from horses by their esterase activity alone.
Ward AC, Sriranganathan N, Evermann JF, Traub-Dargatz JL.Escherichia coli was isolated from the feces and intestines of foals with and without diarrhea. Piliation of isolates was demonstrated by electron microscopy and agglutination in antisera having specificity for K88, K99, P987 and F41 pili. Piliation was also demonstrated by electron microscopy on organisms which did not react with any of the antisera.
Harkness RA, McCreanor GM, Allsop J, Snow DH, Harris RC, Rossdale PO, Ousey JC.1. Plasma hypoxanthine and xanthine concentrations are very low in the horse and low in rat, mouse and greyhound compared to concentrations in beagles, man, sheep and rabbit. 2. Activities in erythrocytes of the main enzyme metabolizing hypoxanthine, hypoxanthine phosphori-bosyltransferase, show a similar pattern (Tax et al., 1976, Comp. Biochem. Physiol. 54B, 209-212); thus low activities have been found where plasma concentrations were low. 3. Hypoxanthine phosphoribosyltransferase activities in horse tissue other than erythrocytes are similar to those in man and rabbit with high activities ...
Klein LV, Soma LR, Nann LE.To investigate the accuracy and precision of the portable, battery-powered StatPal II and the laboratory-based NOVA StatProfile 1 blood gas and pH analyzers for use in analysis of equine blood. Methods: Patient sample comparison and whole blood tonometry. Methods: Patient sample comparison: 125 arterial or venous blood samples from 49 healthy, awake, or anesthetized horses or ponies. Tonometry: venous blood samples from 11 healthy Thoroughbred horses. Methods: Arterial and venous blood taken from awake and anesthetized equine patients was placed in an ice-water bath, then analyzed within 30 mi...
Vliegenthart JF.This research focuses on the study of glycoproteins, specifically investigating their carbohydrate chains and their various functions in living organisms. The article highlights the challenges in isolating specific carbohydrate chains […]
Brobst DF, Carroll RJ, Bayly WM.Urinary concentrations of gamma glutamyl transferase (GGT), alkaline phosphatase (AP), aspartate aminotransferase (AsAT), and alanine aminotransferase (AAT) were measured in 32 healthy horses (16 geldings and 16 females) at the same time on 2 consecutive days. The subjects were divided into 4 ages groups, each comprising 8 horses (4 of each sex). In 10 of the geldings, urine was collected continuously for 72 hours, with catheterized samples being obtained at 0800, 1400 and 2000 hours, and an aliquot of the pooled urine being taken every 24 hours. Urinary enzyme activity was found to be unrelat...
van Galen G, Divers TJ, Savage V, Schott HC, Siwinska N.The aim of this consensus statement is to summarize and appraise scientific evidence and combine this with the clinical experience of a panel of experts to optimize recommendations on how to recognize and manage kidney disease in horses.
Goodrich EL, Webb JL.Previous hematologic and serum biochemistry reference interval (RI) values have been established for donkeys in various geographic regions, life-stages, or for specific donkey breeds. The last extensive investigation establishing RIs for adult donkeys in the United States (U.S.) was published over three decades ago. We aimed to establish updated robust RIs using a reference population of apparently healthy adult donkeys from across the U.S. Standard sized ( = 102), miniature ( = 17), and mammoth ( = 1) donkeys from four different states were enrolled, with 20% of the study population including...
Marshall DE, Gower DB, Silver M, Fowden A, Houghton E.Equine umbilicus was cannulated in utero and a series of cord plasma samples removed for analysis. After steroid extraction and derivatisation, gas chromatographic-mass spectrometric (GC-MS) analysis demonstrated large differences in steroid content between the plasma samples obtained from the umbilical artery and vein, the blood supplies leading to and from the placental surface, respectively. 3Beta-hydroxy-5,7-androstadien-17-one, dehydroepiandrosterone, pregnenolone, 3beta-hydroxy-5alpha-pregnan-20-one, 5-pregnene-3beta,20beta-diol and 5beta-pregnane-3beta,20beta-diol were identified as maj...
Roest HI, de Bruijn CM, Picavet MT, Prins B, Parmentier D, de Zwart GM, Dijkstra YE, van Zijderveld FG.Symptoms, diagnosis and therapy of equine botulism are discussed by the presentation of two detailed reports of horses with neurological symptoms and the results of laboratory investigations over the period 2003-2008 in the Netherlands. In addition a brief summary of the available literature is presented. Prevailing symptoms of botulism in horses include paralysis of the tongue, salvation, dysphagia and paresis and paralysis of the skeletal muscles, as well as signs of colic. Symptoms and prognosis vary with the amount of botulinum neurotoxin (BoNT) involved. For early clinical diagnosis of bo...
Bromiley MW.Before a physical therapy and rehabilitation program is suggested, the end requirement must be considered. All physiotherapeutic machines are subject to laboratory screening. In the United States, the approval of the Food and Drug Administration is required; in the United Kingdom, certification by the National Physics Laboratory has been required by law since January 1996. Laboratory experiments are continually conducted to examine and evaluate the effects on tissues of varied electrical waveforms, low-intensity electrical currents, sound waves, and light rays delivered by a variety of therape...
Metz GE, Lorenzón EN, Serena MS, Corva SG, Panei CJ, Díaz S, Cilli EM, Echeverría MG.A peptide-based indirect ELISA was developed to detect antibodies against Equine arteritis virus (EAV). Two peptides for epitope C of protein GP5 and fragment E of protein M were designed, synthesized, purified and used as antigens either alone or combined. Ninety-two serum samples obtained from the 2010 Equine viral arteritis outbreak, analyzed previously by virus neutralization, were evaluated by the ELISA here developed. The best resolution was obtained using peptide GP5. The analysis of the inter- and intraplate variability showed that the assay was robust. The results allow concluding tha...
de la Calle J, Burba DJ, Ramaswamy CM, Hosgood G, Williams J, LeBlanc C, Moore RM.To compare plasma and synovial fluid endothelin-1 (ET-1) and nitric oxide (NO) concentrations in clinically normal horses and horses with joint disease. Methods: 36 horses with joint disease, and 15 horses without joint disease. Methods: Horses with joint disease were assigned to 1 of the 3 groups (ie, synovitis, degenerative joint disease [DJD], or joint sepsis groups) on the basis of findings on clinical and radiographic examination and synovial fluid analysis. Endothelin-1 and NO concentrations were measured in plasma from blood samples, collected from the jugular vein and ipsilateral cepha...
Pavone S, Mandara MT.The interstitial cells of Cajal (ICC) play a key role in the control of intestinal motility and have been implicated in several human gastrointestinal dysmotility syndromes, in equine grass sickness and in other intestinal disorders where a significant reduction in ICC density was observed. Objective: To investigate the density of ICC in clinically normal horses, ICC c-Kit expression was evaluated by image analysis in order to obtain numerical data. Methods: Intestinal samples from the jejunum to small colon from 5 clinically normal horses were studied. Immunohistochemical labelling of ICC was...
Mizukoshi N, Sakamoto K, Iwata A, Ueda S, Kamada M, Fukusho A.The reverse transcription followed by the polymerase chain reaction (RT-PCR) technique was applied to the detection of African horsesickness virus (AHSV) using primers specific for attenuated AHSV serotype 4 segment 5 (NS1 gene). Total RNA which contains both messenger RNA and genomic dsRNA was extracted by the acid guanidinium-phenol-chloroform method from the AHSV infected Vero cells and was used as templates to optimize the RT-PCR. A pair of primer (NP2-NP32) amplified the product of the expected size from all serotypes of attenuated AHSV when four pairs of primers were tested. Using this p...
Halvarsson P, Tydén E.Gastrointestinal nematode parasites are of major concern for horses, where Strongylus vulgaris is considered the most pathogenic among the Strongylus species. Diagnosis of S. vulgaris infections can be determined with next generation sequencing techniques, which are inherently dependent on reference sequences. The best marker for parasitic nematodes is internal transcribed spacer 2 (ITS2) and we provide the first complete ITS2 sequences from five morphologically identified S. vulgaris and additional sequences from two S. edentatus. These sequences have high similarity to already published part...
Lindgren G, Breen M, Godard S, Bowling A, Murray J, Scavone M, Skow L, Sandberg K, Guérin G, Binns M, Ellegren H.We report fluorescence in-situ hybridization (FISH) and somatic cell hybrid mapping data for 13 different horse genes (ANP, CD2, CLU, CRISP3, CYP17, FGG, IL1RN, IL10, MMP13, PRM1, PTGS2, TNFA and TP53). Primers for PCR amplification of intronic or untranslated regions were designed from horse-specific DNA or mRNA sequences in GenBank. Two different horse bacterial artificial chromosome (BAC) libraries were screened with PCR for clones containing these 13 Type I loci, nine of which were found in the libraries. BAC clones were used as probes in dual colour FISH to confirm their precise chromosom...
Yu J, Han KS, Lee G, Paik MJ, Kim KR.The enantioseparation of pranoprofen after its addition in racemic form into equine plasma and urine was conducted by chiral liquid chromatography-tandem mass spectrometry in selected reaction monitoring mode. The methods for the assay of both enantiomers were linear (r≥0.9943) in the low range from 0.001 to 0.1μg/mL and high range from 0.01 to 1.0μg/mL with good precision (% RSD≤5.6) and accuracy (% RE=-5.3 to 1.9). When racemic pranoprofen was orally administered to four horses at a single dose of 3.1mg/kg, the median plasma concentrations of (R)-pranoprofen were lower than the levels ...
Peglar MT, Nerad TA, Anderson OR.A new species of lobosean amoeba, Stenamoeba polymorpha n. sp., was isolated from the diarrheic stool of a domesticated horse in Great Falls Virginia, U.S. It shares characteristics with the five other described Stenamoeba species. However, electron microscopy revealed S. polymorpha has a substantially thickened cell surface lamina. Under light microscopy, the amoebae had a dynamic polymorphic appearance because hyaloplasm readily formed and resorbed subpseudopodia from any peripheral region of the cell. While in locomotion, the amoebae produced subpseudopodia that led and alternated the dire...
Bonnaire Y, Plou P, Pages N, Boudene C, Jouany JM.A highly sensitive procedure for GC/MS determination of etorphine in horse urine is described. This assay provides both specificity and reliability and is particularly well suited for the confirmation of radioimmunoassay screening procedures usually used for etorphine. After solvent extraction and purifications, the etorphine is characterized as a pentafluoroacetic derivative (PFAA) by using mass fragmentography. The detection limit is 0.1 ng/mL in urine; the coefficient of variation of the estimations is 10.9%. The procedure has been validated after on-field administration of 5 to 90 microgra...
Matassa LC, Woodard D, Leavitt RK, Firby P, Beaumier P.Glycopyrrolate (Robinul) is a quaternary ammonium salt which serves as a respiratory enhancing drug. It is reportedly used in horse racing to improve breathing. Extraction of glycopyrrolate from equine urine employing unique solid-phase extraction techniques gave a residue suitable for liquid chromatography-tandem mass spectrometry (LC-MS-MS) and gas chromatography-mass spectrometry (GC-MS). LC-MS-MS analysis employed an extract derived from 5 ml of urine subjected to cation-exchange chromatography. The daughter ion of m/z 318 monitored in the positive-ion mode was m/z 116. Recovery of glycopy...
Stewart DR, Stabenfeldt GH, Hughes JP.Plasma relaxin concentrations were measured hourly by radio immunoassay in 4 pregnant mares from 11 days before until 4 days after natural foaling. Pre-partum levels ranged from 4 to 7 ng/ml without any surge until the second stage of labour when they increased rapidly to about 11 ng/ml. In 3 of these mares, relaxin activity declined immediately after the expulsion of the placenta and was below detectable levels within 36 h. In the other mare relaxin activity did not fall until after the mechanical removal of the placenta 7 h after foaling. Eight mares were induced to foal by the administratio...
Tozaki T, Hirota K, Mashima S, Tomita M, Mukoyama H.A genomic clone isolated from an equine genomic library probed with an oligonucleotide (CAG)10 showed high sequence similarity to the human F18 gene and was tentatively named equine F18 gene. Because the human F18 gene is expressed in many tissues, we examined whether this equine clone was also expressed in equine tissues. The cDNA encoding equine F18 was obtained by the reverse transcriptase-polymerase chain reaction (RT-PCR) from equine thymus. The nucleotide sequence of the equine F18 cDNA (1940 bp) was determined and contained both the ATG initiation codon and a poly(A) sequence. The cDNA ...
You Y, Proctor RM, Haughan J, Missanelli JR, Robinson MA.Fentanyl, a powerful synthetic mu opioid receptor agonist, is banned in equine sports by the Association of Racing Commissioners International and the Fédération Équestre Internationale. The presence of fentanyl in equine blood has been confirmed during routine post-race screening for doping substances in the authors' laboratory. While fentanyl can be detected and confirmed in blood, it is rapidly metabolized, and screening for the metabolite N-[1-(2-phenethy-4-piperidinyl)] maloanilinic acid (PMA) in equine urine is expected to allow for a longer detection time. In this study, a quantitati...
