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.
Salminen K.The levels of serum vitamin B were determined on 16 mature partly warm-blooded, partly Finnish rural-race horses by the radioisotopic competitive inhibition assay method. The mean value from three samplings carried out in dupli- or triplicate was 1.54 ± 0.16 ng/ml. The utilization of serum inorganic cobalt for cyanocobalamin synthesis was studied on two geldings, which received a dose of 200 µCi CoGl i.v. A Sephadex G-100 gel filtration was carried out with the serum proteins from serial blood samplings at different time intervals 15 min. to 48 hrs. after administration. The gel filtration s...
Milgrom F, Loza U, Kano K.Double diffusion tests in gel were employed for studies of reactions between infectious mononucleosis sera and extracts of bovine, sheep and equine erythrocyte stromata. The extracts were obtained by ultrasonication of stromata prepared from trypsin-digested erythrocytes. The reaction with bovine stroma extract was composed, in many instances, of two lines. A single line was observed in reactions with sheep and equine stroma extracts. This line merged into a reaction of partial or complete identy with one of the lines formed with bovine stroma extract. Evidence was obtained that some infectioo...
Oeding P, Hájek V, Marsálek E.Out of 70 S. aurew strains isolated from the anterior nares of horses, 48 (69 per cent)
belonged to the E biotype. Approximately one third of these isolates were typed with factor
sera, the 6 (35 per cent) that were typable showing 5 different patterns. All strains but one
were non-typable with the basic sets of phages for typing human and bovine staphylococci
even at RTD x 100. Without any exception the equine staphylococci of the E biotype
contained polysaccharide Aa. Sixteen biochemically different strains belonged to the biotype A, B or C. A number of different serological patterns an...
Main AR, Soucie WG, Buxton IL, Arinc E.A relatively simple method is described by which cholinesterase was purified about 19000-fold starting from horse serum. Typically 20 litres of serum were processed to yield 15-18mg of electrophoretically pure cholinesterase in the form of an active salt-free dry powder. The method included two stages: fractionation with (NH(4))(2)SO(4) and ion-exchange chromatography. The (NH(4))(2)SO(4) stage included, in principle, the acid (pH3) step of the Strelitz (1944) procedure. The step took advantage of the stabilizing effect that 33%-satd. (NH(4))(2)SO(4) has on cholinesterase activity at pH3 and i...
Augustinsson KB, Bartfai T, Mannervik B.The steady-state kinetics of the butyrylcholinesterase-catalysed hydrolysis of butyrylthiocholine and thiophenyl acetate were shown to deviate from Michaelis-Menten kinetics. The ;best' empirical rate law was selected by fitting different rate equations to the experimental data by non-linear regression methods. The results were analysed in view of two alternative interpretations: (1) the reaction is catalysed by a mixture of enzymes, or (2) the activity is due to a single enzyme displaying deviations from Michaelis-Menten kinetics. It was concluded that the second alternative applies, and this...
Margalit R, Schejter A.The specific binding of cations to horse heart ferrocytochrome c has been studied, using the gel
filtration method. The cations investigated were: Mg2+, Co2+, cinchonine and proflavine. The stability
constants are in the range of 5-8 mM-1, and the number of binding sites per protein molecule are
1 to 2. The temperature dependence of the stability constant for the Mg2+-ferrocytochrome system
was measured. The thermodynamic parameters were found to be: dH&s = 4-12 kcal/mol, dG;,,
(25 "C) = -5.6 kcal/mol and AS&, = +57 calxmol-lx K-I.
Vaughn DM, Smyth GB.The serotonin metabolite, 5-hydroxyindoleacetic acid (5-HIAA) and the dopamine metabolite, homovanillic acid (HVA) in the cerebrospinal fluid (CSF) of seven clinically normal horses were evaluated with reverse phase high pressure liquid chromatography and electrochemical detection. Comparisons of the neurotransmitter metabolite concentrations were made on CSF collected simultaneously from the atlanto-occipital and lumbosacral regions. There were significantly higher amounts of 5-HIAA and HVA in atlanto-occipital CSF than in lumbosacral CSF. Mean 5-HIAA concentrations in atlanto-occipital and l...
Nelis SA, Sievers C, Jarrett M, Nissen LM, Kirkpatrick CM, Shaw PN.In this paper, a method for the sensitive and reproducible analysis of lignocaine and its four principal metabolites, monoethylxylidide (MEGX), glycylxylidide (GX), 3-hydroxylignocaine (3-HO-LIG), 4-hydroxylignocaine (4-HO-LIG) in equine urine and plasma samples is presented. The method uses liquid chromatography coupled to tandem mass spectrometry operating in electrospray ionisation positive ion mode (+ESI) via multiple reaction monitoring (MRM). Sample preparation involved solid-phase extraction using a mixed-mode phase. The internal standard adopted was lignocaine-d(10). Lignocaine and its...
Benoit E, Jaussaud P, Besse S, Videmann B, Courtot D, Delatour P, Bonnaire Y.A benzhydrolic metabolite of ketoprofen, formed by reduction of the keto group of the drug, has been identified by gas chromatography-mass spectrometry in equine plasma and urine. After partial synthesis, its structure has been confirmed by UV, IR and 1H NMR spectroscopy. The kinetics of ketoprofen and this metabolite have been monitored in plasma by high-performance liquid chromatography. The two products were quantified in plasma up to 4 and 3 h, respectively, and were detected in urine up to 72 and 24 h, respectively, after a single intravenous administration to horses at the dose of 2.2 mg...
Kravtsov AL.Compare the content of bactericidal granules (BG) in blood phagocytes of animals, that differ by species sensitivity to plague infection, under the conditions of measuring, that ensure automatic differentiating by this parameter of monocytes and granulocytes of human blood. Methods: Human whole blood leukocytes were studied, as well as from 7 animal species: mice, guinea pigs, golden hamsters, white rats, rabbits, dogs and horses. Acridine orange (AO) was used for supra-vital staining in primary (bactericidal) granule cells. Relative BG content was measured in separate cells in conventional un...
Petry S, Breuil MF, Duquesne F.Contagious equine metritis (CEM) detection by PCR is recognized by the European Union according to Commission Implementing Regulation (EU) No 846/2014, and real-time PCR is now recommended by the World Organisation for Animal Health Terrestrial Manual at the same level as the culture method. The present study highlights the creation of an efficient network of approved laboratories in France in 2017 for CEM detection by real-time PCR. The network currently consists of 20 laboratories. A first proficiency test (PT) was organized by the national reference laboratory for CEM in 2017 to evaluate th...
George JW, Duncan JR, Mahaffey EA.Pyrimidine 5'nucleotidase (P5N) activities of erythrocytes for Mongolian gerbils, cats, dogs, sheep, horses, and calves were measured, using a radiometric technique with [14C]cytidine monophosphate as the substrate. Erythrocytes of gerbils had the highest activity [1,177.1 +/- 133.6 mU/g of hemoglobin (Hb)]. Feline erythrocytes had 327.4 +/- 204.4 mU/g of Hb. Canine erythrocytes had 148.0 +/- 19.8 mU/g of Hb. Ovine erythrocytes (44.3 +/- 20.9 mU/g of Hb), equine erythrocytes (30.0 +/- 15.9 mU/g of Hb), and bovine erythrocytes (14.1 +/- 6.9) had relatively low P5N activity. The P5N activity was...
Shen Y, Wang WJ, Fu M, Xu GQ, Zhou X, Liu B.To identify the original components of Asini Colla Corii and its raw material hides provides a guarantee for authenticity of Asini Colla Corii. It is urgent for Asini Colla Corii production enterprises and market supervision departments to develop effective identification methods of Asini Colla Corii and hides derived from horses, donkeys, mules and hinnies. This study screened species-specific DNA sequences of nuclear and mitochondrial genomes as detection targets, designed horse and donkey specific primers and established multiple PCR identification methods for identifying the animal hides (...
Kirschvink N, Art T, Lekeux P, Roberts C, Gustin P.8-Epi-PGF2alpha, a prostaglandin-like compound generated by oxidative stress, has been shown to be an in vitro bronchoconstrictor in airways from healthy laboratory animals and healthy humans, but it has never been studied in diseased airways. Here, the bronchoconstrictive capacity of 8-epi-PGF2alpha on isolated bronchial rings (BR) of healthy and heaves-affected horses was evaluated by comparing the maximal effect and the potency of 8-epi-PGF2alpha to those of (1) acetylcholine (ACh), (2) its stereoisomer PGF2alpha and (3) its synthetic receptor agonist, U46619. Furthermore, the potential cap...
Zierz J, Wintzer HJ.To judge acute processes of pain objectively the results are told of a determination of adrenaline and noradrenaline in the plasma of 30 horses suffering from pain. Besides a scheme basing on an awarding of points is developed to ascertain changes of physiological and ethological parameters caused by pain. These results in changes of behaviour are compared to results determined by laboratory experiments. Concerning pain of medium and high level a relation to the concentration of catecholamines is noticed. Therefore the total of certain clinical observations is suitable for graduating acute pai...
Pusterla N, Mapes S, Leutenegger CM.A questionnaire was developed to document the knowledge base of large-animal diplomates of the American College of Veterinary Internal Medicine (ACVIM) regarding polymerase chain reaction (PCR) technology and to identify the common use of this technology in equine practice. Ninety-three of the 278 mailed questionnaires were returned, for an overall response rate of 33.4%. Ninety respondents (99%) reported being familiar with the general principles of nucleic acid probe technology; however, only 52 (57%) knew the difference between conventional (traditional) and real-time (second-generation) PC...
Bridges CG, Edington N.Equine sera were used to immunoprecipitate radiolabelled virus-infected cell proteins; subsequent resolution with polyacrylamide gel electrophoresis identified the EHV-1 polypeptides VP 2, 10a, 11, 13, 14, 15, 16, 20, 21 and 23a. The humoral support of ADCC by these sera was examined in vitro. Cytotoxicity could be demonstrated against both subtypes irrespective of the immunising isolate. The implications of these results are discussed.
Gerdemann R, Deegen E, Kietzmann M, Venner M.In the present study the significance of eicosanoids in the development of shock in horses on the basis of ileus has been investigated using the prostanoids thromboxane B2 (TXB2) and prostaglandine E2 (PGE2) as indicators. The prostanoid synthesis inhibitor flunixin meglumine was to be examined regarding its efficacy in the effective blockade of the synthesis of these mediators within the peri-operative timeframe as well as its effects on clinical signs and laboratory parameters. 21 horses suffering from ileus and ready for surgical intervention received an intravenous flunixin dosis of 1.1 mg...
Constable PD, Tinkler SH, Couëtil LL.To determine the degree of agreement between 2 analyzers for measurement of total CO₂ concentration (ctCO₂) in equine plasma. Methods: 6 healthy untrained horses, 6 trained Standardbreds undergoing a simulated race protocol, and 135 trained Standardbreds at a racetrack. Methods: Jugular venous blood samples were obtained from all horses. Two analyzers (commonly used analyzer A and less expensive analyzer B) were used to measure plasma ctCO₂ in each sample. Validation of both analyzers was conducted in accordance with guidelines established by the Clinical and Laboratory Standards Institu...
Serafini R, Ghosh S, Love CC, Medrano JMR, Teague SR, LaCaze KA, Varner DD.Commercially available vaginal lubricants, typically labeled as non-spermicidal, are used to lubricate equine artificial vaginas prior to semen collection. Improper type or amount of lubricant might affect stallion sperm quality, either after short-time exposure or following cooled storage of extended semen previously exposed to lubricant. The aim of this study was to evaluate stallion sperm quality following exposure to lubricant-containing extender for 1 h (T1h) or 24 h (T24h). Three ejaculates were collected from each of four stallions using a small volume of petrolatum to lubricate a...
Pemberton DH, Thomas KW, Terry MJ.The prevalence of hypogammaglobulinaemia in 82 young foals was determined. Twelve foals were considered clinically abnormal at birth and ten died within two weeks. All of these foals were hypogammaglobulinaemic. Seven (10%) of the other 70 apparently normal foals were hypogammaglobulinaemic despite having suckled normally. Three of these foals developed significant disease and one died at one month of age. Rapid detection of foals with low serum immunoglobulin levels was achieved by adapting the zinc sulphate turbidity test to partially evacuated blood collection tubes. This permitted test to ...
Mogielnicka-Brzozowska M, Woźniak MJ, Cichowska AW, Fraser L, Kraziński B, Strzezek R, Zielińska D.Seminal plasma (SP) proteins are responsible for sperm functional quality. Developing a reliable method to determine the degree of oxidative damage of these proteins is important for establishing semen fertilizing ability. The main aim of the study was to verify the applicability of protein carbonyl derivatives measurement in the SP of canine and stallion, using a method with 2,4-dinitrophenylhydrazine (DNPH). The research material consisted of ejaculates obtained from eight English Springer Spaniels, and from seven half-blood stallions during the breeding and non-breeding season. The content ...
Inoue OJ, Freeman DE, Wallig M.To study effects of hypochlorous acid (HOCl) on equine colonic mucosa in vitro, and determine whether addition of ascorbic acid protects against the effects. Methods: 6 healthy horses and ponies. Methods: Short-circuit current was measured in mucosa mounted in Ussing chambers. Incubation conditions were: control (no additions); 5 mM HOCl; 1 mM HOCl; same and 5 mM ascorbic acid; 3 mM HOCl; 3 mM HOCl and 5 mM ascorbic acid; 7 mM HOCl; and 7 mM HOCl plus 5 mM ascorbic acid. Permeability was measured with [3H]mannitol and, at the conclusion of each experiment, tissues were examined microscopically...
Pu F, McKinney AR, Stenhouse AM, Suann CJ, McLeod MD.A study of the equine phase II metabolism of the anabolic agent boldenone is reported. Boldenone sulfate, boldenone glucuronide and their C17-epimers were synthesised as reference standards in our lab and a method was developed for their detection in a horse urine matrix. Solid phase extraction was used to purify the analytes, which were then detected by ion trap LC/MS. Negative and positive ionisation mode MS(2) were used for the detection of sulfate and glucuronide conjugates, respectively. Boldenone sulfate and 17-epiboldenone glucuronide were detected as the major and minor phase II metabo...
Pihl TH, Jacobsen S, Olsen DT, Højrup P, Grosche A, Freeman DE, Andersen PH, Houen G.OBJECTIVE To purify and characterize equine vitamin D-binding protein (VDBP) from equine serum and to evaluate plasma concentrations of VDBP in healthy horses and horses with gastrointestinal injury or disease. ANIMALS 13 healthy laboratory animals (8 mice and 5 rabbits), 61 healthy horses, 12 horses with experimentally induced intestinal ischemia and reperfusion (IR), and 59 horses with acute gastrointestinal diseases. PROCEDURES VDBP was purified from serum of 2 healthy horses, and recombinant equine VDBP was obtained through a commercial service. Equine VDBP was characterized by mass spectr...
Gu X, Meleka-Boules M, Chen CL.A capillary electrophoresis technique was developed for the separation of synthetic glucocorticoids and the determination of dexamethasone and flumethasone in horse urine. Pretreatment of the sample using a dexamethasone affinity column resulted in low background that enabled the authors to detect levels as low as 1.1 ng/mL and 2.7 ng/mL for dexamethasone and flumethasone in horse urine, respectively. The developed method was used to detect dexamethasone in horse urine samples after the injection of a therapeutic dose of dexamethasone for up to 12 hr postinjection. The optimum conditions for c...
Kaneps AJ, Craig AM, Walker KC, True JE.To determine whether iontophoretic administration of dexamethasone to horses results in detectable concentrations in synovial fluid, plasma, and urine. Methods: 6 adult mares. Methods: Iontophoresis was used to administer dexamethasone. Treatments (4 mA for 20 minutes) were administered to a tarsocrural joint of each mare. The drug electrode contained 3 ml of dexamethasone sodium phosphate at a concentration of 4 or 10 mg/ml. Samples of synovial fluid, blood, and urine were obtained before and 0.5, 4, 8, and 24 hours after each treatment. All samples were tested for dexamethasone using an ELIS...
Legrottaglie R, Agrimi P.Electrophoretic analysis in polyacrylamide gel (PAGE) of the equine rotavirus 106/88/LI/EQ, isolated from the diarrhea of an 18 day old foal was compared to the bovine strain NCDV. There was a notable difference in the migration of some segments of the viral RNA. Bands 2 and 3 of the equine rotavirus comigrated while there was a clear separation of segments 7, 8 and 9. Moreover, the migration of segments 1, 4 and 5 revealed a lower molecular weight than the corresponding segments of NCDV.
Thompson DL, McNeill DR, Wiest JJ, St George RL, Jones LS, Garza F.Sequential samples of blood were drawn via jugular catheters every 15 min for 24 h from four mares in each of five reproductive states: intact anestrous mares in winter, intact diestrous mares in summer, intact estrous mares in summer, ovariectomized mares in winter and ovariectomized mares in summer. Estrous mares were sampled on d 4 or 5 of estrus and diestrous mares on d 10 or 11 of diestrus. Each sample of plasma was assessed for concentrations of luteinizing hormone (LH) and follicle stimulating hormone (FSH) in two independent radioimmunoassays. A computer program was developed that dete...
Morris CM, Field HJ.Tissue specimens obtained from equine herpesvirus-1 (EHV-1), subtype 1-infected aborted foetuses were analysed for the presence of virus DNA by means of Southern blot and dot blot hybridisations. The specificity of the methods was confirmed although the sensitivity was inferior to classical techniques such as virus isolation. However, the possibility of detecting the state of the virus DNA and the ability to distinguish between subtypes were important features, and the dot blot method was shown to have potential for a rapid diagnostic test. This report demonstrates some potential practical app...
Wong JKY, Leung DKK, Curl P, Schiff PJ, Lam KKH, Wan TSM.The detection of boldenone, nandrolone, 5(10)-estrene-3β,17α-diol, and 4-estrene-3,17-dione in a urine sample collected from a gelding having been treated with testosterone (500 mg 'Testosterone Suspension 100', single dose, injected intramuscularly) in 2009 led the authors' laboratory to suspect that these 'testicular' steroids could be minor metabolites of testosterone in geldings. Administration trials on six castrated horses with Testosterone Suspension 100 confirmed that low levels of boldenone, nandrolone, 5(10)-estrene-3β,17α-diol, and 4-estrene-3,17-dione could indeed be detected ...
Ainsworth DM, Dodds WJ, Brown CM.A 16-year-old gelding was examined because of weight loss, inappetence, and intermittent fever of 2 months' duration. Preliminary laboratory findings revealed anemia, hypoproteinemia, thrombocytopenia, and prolongation of the activated partial thromboplastin time. A deficiency or inhibition of coagulation factor XI, factor XII, or high molecular weight kininogen was diagnosed. This defect was not associated with a bleeding diathesis, but should be considered as a cause of prolongation of the activated partial thromboplastin time.
Morris DD, Crowe N, Moore JN.The purpose of this study was to determine if a structurally novel dual inhibitor of arachidonic acid metabolism, SK & F 86002, would inhibit the endotoxin-induced production of tumor necrosis factor (TNF) activity by equine peritoneal macrophages. Equine peritoneal macrophages were variously pretreated for 0, 0.5 and 2 h with SK & F 86002 at 10(-9) to 10(-4) molar final concentrations or were left untreated. Then, the macrophages were cultured in vitro in the presence of endotoxin (5 ng/mL). Supernatant media were collected after 4 h and stored at -70 degrees C until assayed for TNF a...
Delbeke FT, Baert K, De Backer P.Urinary and plasma concentrations of the nonsteroidal anti-inflammatory drug tiaprofenic acid were determined following oral and intramuscular administration of a dose of 1 g to five fasted horses. Quantitation was performed by high-performance liquid chromatography (HPLC). The limit of quantitation (LOQ) was 0.1 microg/ml and 0.5 microg/ml in 2 ml plasma and 1 ml urine, respectively. Assay precision and extraction recovery were between acceptable values. Tiaprofenic acid pharmacokinetics were described by non-compartment analysis of the data. Absorption was faster after oral administration as...
Whan G, Phillips J, Bullock S, Runciman RJ, Pearce S, Hurtig M.A novel, multi-use, low-stiffness and low-cost transducer for measuring in vitro strains has been developed and tested. Currently available strain measurement methods are either too expensive, too complicated or too inflexible for multi-use strain measurement. The stainless-steel modular strain measurement clip introduced here was instrumented with four 350 Omega axial strain gauges in a full Wheatstone bridge configuration to take advantage of commonly available strain gauge amplifier equipment. Adjustable extension arms were designed to allow greater application versatility. The clip was cal...
Carstanjen B, Amory H, Youssao I, Remy B.The purpose of this study was to validate an equine-specific osteocalcin (OC) radioimmunoassay (RIA) for use in donkeys and to establish age-related changes in serum OC concentrations in healthy donkeys. Serial dilutions of donkey serum showed parallelism with standard curves obtained with the equine-specific OC RIA. There was a tight linear regression between donkey serum OC values obtained with the equine specific OC RIA and a commercially available bovine-specific OC RIA. Serum OC levels of 27 healthy donkeys, analysed with the equine-specific OC RIA, showed a tight negative logarithmic reg...
