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.
Cheadle MA, Lindsay DS, Rowe S, Dykstra CC, Williams MA, Spencer JA, Toivio-Kinnucan MA, Lenz SD, Newton JC, Rolsma MD, Blagburn BL.An IFAT was used to determine the prevalence of Neospora-specific IgG antibodies in serum from Alabama horses. Serum samples (n = 536) were from asymptomatic horses routinely submitted for equine infectious anaemia virus infection testing. We also subjected a 13-year-old horse with CNS disease to necropsy examination for isolation and in vitro cultivation of protozoal organisms. In antemortem tests, this horse was positive for antibodies to Neospora sp. in the IFAT and western immunoblot. Results of the prevalence survey indicated that IgG antibodies to Neospora were present in 62 (11.5%) of t...
Cerón JJ, Tecles F, Espín JC.Effects of seven different blood diluents (distilled water, Triton X-100, saponin, isotonic saline solution, pH 7.5 and 8 phosphate buffers and bovine serum albumin) and two chromophores: 5, 5'-dithiobis 2-nitrobenzoic acid (DTNB) and 2,2'-dithiodipyridine (2- PDS) on blood cholinesterase determination in four domestic species (cow, sheep, goat and horse) are described and compared. Haemolytic diluents (distilled water, Triton X-100 and saponin) gave the best precision results when fresh blood was assayed. However, Triton X-100 induced lower ChE activity values in horses, and saponin yielded v...
Todi F, Mendonca M, Ryan M, Herskovits P.The origin of caffeine detections in equine serum and urine after theophylline administrations was examined. Three different preparations containing theophylline were administered to standardbred mares. Both blood and urine samples were collected. Caffeine was detected and quantified in theophylline administration samples by high performance liquid chromatography (HPLC) and liquid chromatography-tandem mass spectrometry (LC-MS-MS). Further in vitro analysis showed that caffeine metabolites were not detected when caffeine, or caffeine-containing products, were added to urine. Data derived from ...
Sternberg S, Johannisson A, Magnusson U, Jensen-Waern M.After exposure of equine granulocytes from both foals and adult horses to culture supernatants from clinical isolates of Actinobacillus equuli, phagocytic capacity and respiratory burst was examined by flow-cytometry and a chemiluminescence assay, respectively. One haemolytic isolate of an equine Actinobacillus was also included in the study. An average decrease of 22% in total number of granulocytes, in the flow cytometric assay (P < 0.01), and an average decrease of 26% in light emission, in the chemiluminescence assay (P < 0.001), was seen after exposure to bacterial culture supernata...
Leutenegger CM, von Rechenberg B, Huder JB, Zlinsky K, Mislin C, Akens MK, Auer J, Lutz H.Specific amplification and quantitation of nucleic acid sequences by the polymerase chain reaction (PCR) has been extensively used for the detection of viral infection and gene expression. Although successful amplification of DNA and RNA sequences extracted from paraffin embedded tissue have been described, there are presently no reports available regarding RNA analysis from bone and calcified tissues embedded in hydrophobic acrylic resin. Here we describe a general method for quantitation of specific mRNA sequences extracted from undecalcified bone sections, fixed in paraformaldehyde, and emb...
Li F, Leroux C, Craigo JK, Cook SJ, Issel CJ, Montelaro RC.Equine infectious anemia virus (EIAV) is genetically one of the simplest lentiviruses in that the viral genome encodes only three accessory genes, tat, rev, and S2. Although serological analyses demonstrate the expression of the S2 protein in persistently infected horses, the role of this viral gene remains undefined. We recently reported that the S2 gene is not essential for EIAV replication in primary equine macrophages, as EIAV mutants lacking the S2 gene replicate to levels similar to those of the parental virus (F. Li, B. A. Puffer, and R. C. Montelaro, J. Virol. 72:8344-8348, 1998). We n...
Idrissi Bougrine S, Fassi Fihri O, el Harrak M, Fassi Fehri MM.A vaccination protocol involving three horses, with five repeated injections of inactivated serotype 4 African horse sickness virus, was undertaken to determine a possible threshold for the appearance of antibodies against the non-structural protein NS3. Using an indirect enzyme-linked immunosorbent assay, with the recombinant NS3 protein as an antigen, the authors detected a response to NS3 as of the second injection for the first horse and after four injections for the second horse. No response to NS3 was detected for the third horse. The results show that the inactivated vaccine is insuffic...
Yachida Y, Kashiwagi M, Mikami T, Tsuchihashi K, Daino T, Akino T, Gasa S.A novel galactosylalkylglycerol modified with a long-chain cyclic acetal at the sugar moiety, 3-O-(4'6'-plasmalogalactosyl) 1-O-alkylglycerol, was isolated from equine brain. The presence of cyclic acetal linkage, its linked position, and the length of the acetal chain of the natural plasmalo lipid were determined by proton NMR spectroscopy and fast-atom bombardment;-mass spectrometry, as well as gas chromatography;-mass spectrometry and gas;-liquid chromatography. To identify the isomeric stereostructure of the natural product, the plasmalo derivative was chemically synthesized from 3-O-galac...
Guérin G, Bailey E, Bernoco D, Anderson I, Antczak DF, Bell K, Binns MM, Bowling AT, Brandon R, Cholewinski G, Cothran EG, Ellegren H, Förster M....The goal of the First International Equine Gene Mapping Workshop, held in 1995, was the construction of a low density, male linkage map for the horse. For this purpose, the International Horse Reference Family Panel (IHRFP) was established, consisting of 12 paternal half-sib families with 448 half-sib offspring provided by 10 laboratories. Blood samples were collected and DNA extracted in each laboratory and sent to the Lexington laboratory (KY, USA) for dispatch in aliquots to 14 typing laboratories. In total, 161 markers (144 microsatellites, seven blood groups and 10 proteins) were tested f...
Murphy AJ, Mansfield LS.Sarcocystis neurona is a protozoan parasite that causes a neurological disease in horses called equine protozoal myeloencephalitis. The route of transmission is speculated to be by fecal-oral transfer of sporocysts shed from opossums. Controversy exists regarding both the natural life cycle for this parasite as well as the species identity of opossum Sarcocystis. To provide stage-specific material for species comparison, 27 opossums from southern Michigan were screened for Sarcocystis spp. sporocysts. Seven opossums were positive for Sarcocystis sporocysts by fecal flotation. A simplified, eff...
Gleadhill A, Marlin D, Harris PA, Michell AR.Measurement of renal function in horses poses a particular challenge because plasma creatinine is influenced by muscle mass which is highly developed and variable between individuals, while conventional clearance methods involve potentially daunting problems, particularly urine collection and bladder washout. This paper provides data which enable technetium-diethyleneaminopentacetic acid (Tc-DTPA) clearance to be used to calculate glomerular filtration rate (GFR)/extracellular fluid volume (ECFV) as an expression of GFR in horses, as previously validated in humans, dogs and calves. Apart from ...
Nagaoka K, Nambo Y, Nagamine N, Nagata SI, Tanaka Y, Shinbo H, Tsunoda N, Taniyama H, Watanabe G, Groome NP, Taya K.The relationship between a selective increase in circulating immunoreactive (ir)-inhibin and the time of ovulation was investigated in mares. Concentrations of plasma ir-inhibin were measured every 4 h during the periovulatory period. Inhibin pro-alphaC, a precursor protein of the inhibin alpha-subunit, was also measured. The changes in ir-inhibin and inhibin pro-alphaC in circulation were parallel. Concentrations of both ir-inhibin and inhibin pro-alphaC in the plasma increased at the same time when ovulatory follicles ruptured, and the peak levels of circulating ir-inhibin and inhibin pro-al...
Schroeder B, Boyle MD, Sheerin BR, Asbury AC, Lottenberg R.Our laboratory previously demonstrated that group C streptococcal isolates from humans and horses secrete streptokinases that preferentially activate plasminogens reflecting the origin of the isolates. To analyze the significance of these findings, series of streptokinase-producing Streptococcus equisimilis isolates recovered from humans and horses were examined. Southern blot analysis revealed that chromosomal DNA of the streptococcal isolates from humans reacted exclusively with a skc(hu) probe and that chromosomal DNA of streptococcal isolates from horses reacted preferentially with an skc(...
Huang Z, Depoortere I, De Clercq P, Peeters T.Motilin is involved in the regulation of the fasting motility pattern in man and in dog, but may have a different role in other species. Immunoreactive motilin has been demonstrated in several species, but the sequence is mostly unknown. The aim of this study was to isolate and sequence the cDNA encoding the motilin precursor from several mammalian species and from chicken. Total RNA was isolated from the duodenal mucosa of the chicken, dog, cow and horse. In each case single stranded cDNA was synthesized. Motilin cDNA fragments were amplified by PCR, ligated into a plasmid and cloned. Clones ...
Collins MD, Rodriguez Jovita M, Lawson PA, Falsen E, Foster G.Two strains of an unknown Gram-positive, catalase-negative, facultatively anaerobic coccus originating from the reproductive tract of horses were characterized by phenotypic and molecular taxonomic methods. Comparative 16S rRNA gene sequencing studies demonstrated that the two strains constitute a new subline within the lactic-acid group of bacteria, close to, but distinct from, Abiotrophia defectiva, Globicatella sanguinis and close relatives. The unknown bacterium was readily distinguished from other described Gram-positive, catalase-negative cocci by biochemical tests and electrophoretic an...
Caetano AR, Lyons LA, Laughlin TF, O'Brien SJ, Murray JD, Bowling AT.Comparative anchor tagged sequences (CATS) from human Chromosome 5 (HSA5) were used as PCR primers to produce molecular markers for synteny mapping in the horse. Primer sets for 21 genes yielded eight horse-specific markers, which were mapped with the UC Davis horse-mouse somatic cell hybrid panel into two synteny groups: UCD14 and UCD21. These data, in conjunction with earlier human chromosome painting studies of the horse karyotype and synteny mapping of horse microsatellite markers physically mapped by FISH, confirm the assignment of UCD21 to ECA21 and suggest that UCD14 is located on ECA14...
Green MK, Johnston MV, Larsen BS.To elucidate the role of high mass accuracy in mass spectrometric peptide mapping and database searching, selected proteins were subjected to tryptic digestion and the resulting mixtures were analyzed by electrospray ionization on a 7 Tesla Fourier transform mass spectrometer with a mass accuracy of 1 ppm. Two extreme cases were examined in detail: equine apomyoglobin, which digested easily and gave very few spurious masses, and bovine alpha-lactalbumin, which under the conditions used, gave many spurious masses. The effectiveness of accurate mass measurements in minimizing false protein match...
Walter I, Nowotny N.Effects of infection with two different strains of equine herpes virus type 1 (EHV-1; Piber 178/83, Kentucky D) on the cytoskeleton of Vero cells were investigated immunohistochemically, and evaluated by confocal laser scanning microscopy. Twenty four hours post EHV-1 infection the assembly of the microtubulus system of Vero cells was heavily disturbed. The Golgi region was dispersed into vesicles spread throughout the cytoplasm as demonstrated by WGA lectin binding. Other cytoskeletal elements such as cytokeratin, vimentin, and filamentous actin (F-actin) were not affected by EHV-1 infection....
Marr KA, Lees P, Cunningham FM.Adherence to vascular endothelium and extracellular matrix proteins is a pre-requisite for neutrophil accumulation at sites of inflammation. In this study, equine neutrophil adherence to fibronectin and autologous serum-coated plastic in response to PAF, hrIL-8, hrC5a and PMA has been measured. In addition, the mechanisms involved have been investigated using monoclonal antibodies (MoAbs) against the beta2 integrin CD18. PAF and hrC5a caused similar, concentration dependent, increases in adherence to fibronectin- and serum-coated plastic (maximum responses 19 +/- 4% and 19 +/- 3% for PAF and 1...
Auvray P, Sourdaine P, Moslemi S, Séralini GE, Sonnet P, Enguehard C, Guillon J, Dallemagne P, Bureau R, Rault S.In this study, we describe the synthesis of a new family of indolizinone derivatives designed to fit an extrahydrophobic pocket within the active site of aromatase and to strongly inhibit human aromatase. This could help improve the specificity of the inhibitors. Equine aromatase, very well characterized biochemically, is used as a comparative model. Indeed, in a previous comparison between both human and equine aromatases, we described the importance of the interaction between the inhibitor and this pocket for the indane derivative MR 20814. MR 20492 and MR 20494 are more potent inhibitors of...
Yokoyama S, Radlwimmer FB.To elucidate the molecular mechanisms of red-green color vision in mammals, we have cloned and sequenced the red and green opsin cDNAs of cat (Felis catus), horse (Equus caballus), gray squirrel (Sciurus carolinensis), white-tailed deer (Odocoileus virginianus), and guinea pig (Cavia porcellus). These opsins were expressed in COS1 cells and reconstituted with 11-cis-retinal. The purified visual pigments of the cat, horse, squirrel, deer, and guinea pig have lambdamax values at 553, 545, 532, 531, and 516 nm, respectively, which are precise to within +/-1 nm. We also regenerated the "true" red ...
Chapman MR, Kearney MT, Klei TR.With the increased interest in equine cyathostomes it has become apparent that some evaluations of methods currently used to count the various larval stages which occur in the mucosa would be beneficial. Experiments were conducted to investigate the effects of fixation and storage of mucosal tissues at -20 C on the accuracy of counting these larvae. The accuracy of counting developing larvae within the mucosa by transmural illumination (TMI) and by artificial digestion (DIG) of the mucosa was also compared. The data indicate that fixation of digested mucosa in PBS-buffered 5% or 10% formalin d...
Neubauer A, Meindl A, Osterrieder N.The equine herpesvirus 1 (EHV-1) modified live vaccine strain RacH is apathogenic for both laboratory animals and the natural host. The apathogenicity of RacH was caused by serial passages of the virus in heterologous cells. When compared to the virulent parental strain RacL11 several changes in the RacH genome occurred. Previous results have shown that the loss of the IR6 gene correlated with the loss of virulence. Additional important mutations were observed within the US2 gene which is directly adjacent to the IR6 gene and within the glycoprotein B (gB) gene. To answer the question whether ...
Sams RA, Gerken DF, Ashcraft SM.The objectives of the study were to compare various methods to determine flunixin in test samples collected periodically from horses after intramuscular (IM) and intravenous (IV) dosing at the maximum recommended dosage and to document detection times for this drug in test samples. Flunixin, a nonsteroidal anti-inflammatory drug approved for use in horses, was administered to eight mares in five consecutive daily doses of 1.1 mg per kilogram of body weight by the IM or IV route. Flunixin was detected in urine samples collected at various times after drug administration by flunixin enzyme-linke...
Wilson SM, Gallagher M, Rakhit S, Remsbury AL, Ko WH.Uridine triphosphate (UTP) evoked inhibition of adrenaline-evoked cAMP accumulation in cultured equine epithelial cells (EC50, 1.8 +/- 0.2 microM) and this effect was mimicked by 5-Br-UTP (EC50, 6.6 +/- 1.8 microM) and uridine diphosphate (UDP; EC50, 96 +/- 26 microM). This inhibitory action of UTP was abolished by pre-treating cells with pertussis toxin (10 ng ml-1, 24 h). UTP (EC50, 2.3 +/- 0.3 microM) and 5-Br-UTP (EC50, 29.4 +/- 9.4 microM) also increased intracellular free calcium ([Ca2+]i) whilst UDP did not; the two effects are thus differentially sensitive to these pyrimidine nucleotid...
Nasir L, Reid SW.The sarcoid is a benign locally invasive dermal fibroblastic lesion, commonly affecting horses and donkeys. The aetiology of the equine sarcoid is equivocal. Bovine papillomaviral (BPV) DNA (type 1/2) is frequently demonstrable in equine sarcoid tumour biopsies. However, the exact role of the virus in the disease process and its contribution to the phenotypic differences in sarcoids is not known. It was sought to assess the transcriptional activity of BPV-1 found in sarcoid tissues. Of 20 tumours examined, 18 were positive for E2 expression and ten positive for L1 expression. Viral oncogenes E...
Komar N, Dohm DJ, Turell MJ, Spielman A.To determine whether eastern equine encephalitis (EEE) virus infection in starlings may be more fulminant than in various native candidate reservoir birds, we compared their respective intensities and durations of viremia. Viremias are more intense and longer lasting in starlings than in robins and other birds. Starlings frequently die as their viremia begins to wane; other birds generally survive. Various Aedes as well as Culiseta melanura mosquitoes can acquire EEE viral infection from infected starlings under laboratory conditions. The reservoir competence of a bird is described as the prod...
Henderson K, Stevens S, Bailey C, Hall G, Stewart J, Wards R.The relative merits of measuring blood concentrations of equine chorionic gonadotrophin (eCG, previously known as pregnant mare serum gonadotrophin (PMSG)), or oestrone sulphate (OS), or faecal OS concentrations for determining pregnancy status in miniature horses were investigated. Pregnant mares between 40 and 140 days after mating had serum eCG concentrations > 1 I.U. mL-1, with the highest concentrations occurring between days 50 and 120. However, eCG measurements were susceptible to returning a 'false positive' diagnosis of pregnancy. Plasma OS concentrations ranged from 0.1 to 3.6 ng ...
Piper T, Geyer H, Gougoulidis V, Flenker U, Schänzer W.Boldenone (androsta-1,4-dien-17beta-ol-3-one, Bo) is an anabolic steroid known to have been used in cattle breeding or equine sport as a doping agent for many years. Although not clinically approved for human application, Bo or its main metabolite 5beta-androst-1-en-17beta-ol-3-one (BM1) were detected in several doping control samples. For more than 15 years the possibility of endogenous Bo production in human beings has been discussed. This is a challenging issue for doping control laboratories as Bo belongs to the list of prohibited substances of the World Anti-Doping Agency and therefore th...
Driessen B, Zarucco L, Steffey EP, McCullough C, Del Piero F, Melton L, Puschner B, Stover SM.The volatile anaesthetic sevoflurane is degraded to fluoride (F-) and a vinyl ether (Compound A), which have the potential to harm kidney and liver. Whether renal and hepatic injuries can occur in horses is unknown. Cardiopulmonary, biochemical and histopathological changes were studied in six healthy thoroughbred horses undergoing 18 h of low-flow sevoflurane anaesthesia. Serum F- concentrations were measured and clinical laboratory tests performed to assess hepatic and renal function before and during anaesthesia. Necropsy specimens of kidney and liver were harvested for microscopic examinat...
Bageshlooyafshar B, Vakilian S, Kehtari M, Eslami-Arshaghi T, Rafeie F, Ramezanifard R, Rahchamani R, Mohammadi-Sangcheshmeh A, Mostafaloo Y....In current study we aimed to coat the PLLA scaffold with zinc (Zn) silicate mineral nanoparticles. Then, using equine adipose-derived stem cells (ASCs) we intended to compare the osteogenic induction potency of Zn silicate mineral-coated PLLA scaffold with uncoated PLLA scaffold and tissue culture plastic (TCPS). Adipose tissues were collected from 3 horses, and isolation of ASCs was achieved by enzymatic digestion. PLLA scaffold was successfully prepared using a phase separation method and coated with Zn silicate mineral nanoparticles. The coating efficiency was then characterized by scanning...
Choi YH, Gibbons JR, Canesin HS, Hinrichs K.Prospective studies were conducted to help define procedural factors affecting in vitro embryo production via intracytoplasmic sperm injection (ICSI) of equine oocytes. In experiment 1, use of 10% fetal bovine serum as a protein source in embryo culture medium resulted in a higher blastocyst rate than did use of a combination of 3% fetal bovine serum, 3% equine preovulatory follicular fluid, and 4% human serum substitute (37% vs. 15%, respectively, P < 0.05). In experiment 2, the effect of zinc supplementation (0, 0.5, 1, or 1.5 μg/mL) during IVM was examined. There were no significant di...
Carraway KL, Colton DG, Shin BC, Triplett RB.Bovine and equine erythrocytes have been studied by three different surface modification techniques to investigate the accessibility of the surface components to the external medium. Lactoperoxidase labeling of equine erythrocytes results in a significant labeling of only one membrane component, a 100 000-mol.wt polypeptide corresponding to the membrane-spanning Component III of human erythrocytes. The major sialoglycoprotein of the equine erythrocyte is not labeled. This is in contradistinction to the situation for human and bovine cells, where both components are labeled. The equine membrane...
Leisinger CA, Klein C, Markle ML, Premanandan C, Sones JL, Pinto CRF, Paccamonti DL.Aluteal cycles were induced in the mare to evaluate the effects of progesterone deprivation on the gene expression of embryos and endometrium collected eight days after ovulation. We hypothesized that the transcript expression would be altered during induced aluteal (AL) cycles (low progesterone <1 ng/mL) when compared with control cycles during diestrus (high progesterone; > 4 ng/mL) for 1) the embryonic expression of progesterone-mediated transcripts and those related to normal embryo growth and development and 2) the endometrial expression of progesterone-mediated transcripts and those ...
Coutinho da Silva MA, Seidel GE, Squires EL, Graham JK, Carnevale EM.The effects of semen extender components on the ability of stallion sperm to bind to the zona pellucida (ZP) and the suitability of using bovine ZP for a ZP-binding assay for stallion sperm were investigated in a series of experiments. In Experiment I, binding of stallion sperm to both bovine and equine ZP was significantly increased when a skim milk-based extender (EZM) was used. In Experiment II, a threefold increase in sperm binding to ZP was observed when sperm were diluted in EZM compared with diluents, which contained no milk (TALP, LAC, and EmCare). In Experiment III, centrifuging the s...
Patterson SD, Bell K.Twelve equine protease inhibitory alleles, PiE, H, J, K, L2, O, P, Q, R, V, X, Z, have been characterized in terms of isoelectric point, molecular mass and inhibitory activity to bovine trypsin and chymotrypsin by ISO-DALT electrophoresis. Protein maps for 20 Pi alleles including those of the eight 'Thoroughbred' alleles (PiF, G, I, L, N, S1, S2, U) have now been determined. Five pairs of alleles, S1/S2, G/K, L/L2, P/R and U/Z, possessed varying numbers of common proteins ranging from one protein in the case of G/K and L/L2 to six in the case of U/Z. Based on these results and studies of the a...
Romo-Sáenz CI, Tamez-Guerra P, Olivas-Holguin A, Ramos-Zayas Y, Obregón-Macías N, González-Ochoa G, Zavala-Díaz de la Serna FJ....Equine infectious anemia (EIA) is a highly infectious disease in members of the family, caused by equine infectious anemia virus (EIAV). The disease severity ranges from subclinical to acute or chronic, and causes significant economic losses in the equine industry worldwide. Serologic tests for detection of EIAV infection have some concerns given the prolonged seroconversion time. Therefore, molecular methods are needed to improve surveillance programs for this disease. We attempted detection of EIAV in 6 clinical and 42 non-clinical horses in Nuevo Leon State, Mexico, using the agar gel immu...
Haase F, Siewert C, von Rautenfeld DB, Fischbach JU, Seifert H.Measuring the leg volume of horses is useful for diagnostic and treatment of different diseases in relation to swollen legs. In the present study different methods to quantify the limb volume are compared, the water displacement method, an optoelectronic 2-dimensional body scanner, the so-called perometer and calculation of volume by applying the disc model. As reference method the golden standard of human medicine - the water displacement method was used. We took volume measurements from different tall horses in a defined section of the forelegs between the coronary band and the carpal joint....
Chen CH, Albers JJ.The abilities of apolipoprotein A-I species isolated from humans, horses, sheep, goats and rabbits to activate purified human lecithin-cholesterol acyltransferase and the enzyme from homologous plasmas and plasma of other mammalian species were compared. Each purified apolipoprotein A-I species was individually incorporated into phosphatidylcholine/cholesterol vesicles by the cholate dialysis method to form proteoliposome common substrates (apolipoprotein A-I/phosphatidylcholine/cholesterol molar ratio of 1:250:12.5) for the enzyme activity assay. All apolipoprotein A-I species tested had the ...
Tkachenko AG, Winston GW.Preincubation of horse liver alcohol dehydrogenase (HLADH) with the oxidative agent, tert-butyl hydroperoxide (tBOOH) results in a twofold stimulation of the ethanol dehydrogenase activity of this enzyme. This stimulation was dependent on tBOOH concentration up to 100 mM; above this concentration tBOOH did not further stimulate ethanol oxidation by HLADH. Active-site-directed reagents and classical ADH binary complexes were used to probe the possible mechanism of this activating effect. The rate and extent of stimulation by tBOOH is strongly reduced by binary complexes with NAD(+) or NADH, who...
Main AR.The kinetic and structural relationships of eight electrophoretically pure mammalian serum and liver serine carboxylesterases (CE) and cholinesterases (ChE) have been studied. Eight CE's and ChE's, which were fully resolved but only partially purified, provided additional information. Five of the electrophoretically pure esterases were monomeric, and of these, four belonged to a new and widely distributed class. These four monomeric esterases hydrolyzed choline esters, but at widely differing rates. Thus two were termed monomeric butyrylcholinesterases, mBuChE I and II, and two were monomeric ...
Chen CL, Zhu D, Gillis KD, Meleka-Boules M.To develop a simple and sensitive ELISA for detection of dexamethasone in horse serum and urine. Methods: Blood and urine samples from 3 thoroughbred mares. Methods: A dexamethasone oxime was prepared and conjugated to hemocyanin, bovine serum albumin and to horseradish peroxidase. One- and two-step double-antibody ELISA methods, as well as a radioimmunoassay method, were performed. The one-step ELISA was used to test urine from 3 Thoroughbred mares injected with 5 mg of dexamethasone, IV. Results: The ELISA could detect dexamethasone in the range of 0.01 to 50 ng/ml, with intra- and interassa...
Popot MA, Lacabaratz E, Garcia P, Laroute V, Bonnaire Y, Toutain PL, Cowan DA.The cortisol threshold concentration of 1.0 microg/ml in horse urine adopted by the International Federation of the racing Authorities in 1994 is specific. However, an increase in the sensitivity for the detection of cortisol administration would be helpful. Previous studies have shown that 20beta-dihydrocortisol concentration in urine would be a good indicator of cortisol administration. The purpose of the present work was to estimate the population parameters and the critical values of 20beta-dihydrocortisol and 20beta-dihydrocortisone concentration in urine compared with that of cortisol. U...
Kuhnert-Paul Y, Schmäschke R, Daugschies A.Results of parasitological examination of faecal aliquots may vary between diagnostic laboratories. To examine whether inhomogeneous distribution of worm eggs in faecal samples is responsible for this observation, horse faeces provided for routine diagnosis of helminth infection were examined. Distribution of worm eggs was assessed by examining aliquots taken from different locations of the faecal sample by a combined sedimentation-flotation method (KSFV). In addition, it was tested, whether the homogenization of a larger amount (minimum of 40 g) of faeces before performing KSFV improved repro...
Bailey DN.The relative binding of acetaminophen, lidocaine, phenobarbital, procainamide, quinidine, and theophylline to sera of seven mammalian species was studied. Pooled commercial sera from cow, goat, horse, human, pig, rabbit, and sheep were supplemented with 5 and 10 mM concentrations of each drug. For each serum, each drug, and each drug concentration, equilibrium dialysis was performed in duplicate against phosphate buffer (pH 7.4, 0.1 M, 4 degrees C). Percent drug bound to serum was calculated. Phenobarbital demonstrated more than 20% binding to goat, horse, human, and sheep serum at both 5 and ...
Weiss DJ, Evanson OA.To determine the potential usefulness of tests for detection of platelet activation and platelet-leukocyte aggregates in horses. Methods: Blood from 3 healthy Thoroughbreds. Methods: Microscopic and flow cytometric assays were used to evaluate spontaneous platelet aggregation, platelet activation, and platelet-leukocyte aggregates. Platelet activation was detected by evaluation of binding of anti-human fibrinogen to unactivated and ADP-, thrombin-, thrombin agonist receptor peptide-, and platelet activating factor-activated platelets. Platelet-leukocyte aggregates were evaluated microscopicall...
Zhang P, Sun Y, Tan C, Ling M, Li X, Wang W, Cong Y.To prevent and control H3N8 subtype equine influenza, we prepared virus-like particles (VLPs) comprising the HA, NA and M1 proteins of H3N8 equine influenza virus (EIV) through the insect cell-baculovirus expression system. The results of Western blot and hemagglutination analyses demonstrated that the constructed VLPs comprising HA, NA and M1 proteins have good hemagglutination activity. Immunoelectron microscope revealed that the VLPs share similar morphology and structure with natural virus particles. The hyperimmune serum from horses immunized with the VLPs were injected into mice by means...
Azab W, El-Sheikh A.Equine herpesvirus 4 (EHV-4) is an important equine pathogen that causes respiratory tract disease among horses worldwide. Glycoprotein K (gK) homologues have been identified in several alphaherpesviruses as a major player in virus entry, replication, and spread. In the present study, EHV-4 gK-deletion mutant has been generated by using bacterial artificial chromosome technology and Red mutagenesis to investigate the role of gK in EHV-4 replication. Our findings reported here show that gK is essential for virus replication in vitro and that the gK-negative strain was not able to be reconstitut...
Brink P, Wright JC, Schumacher J.The glutaraldehyde test (GT), a rapid and inexpensive test, has been utilized empirically for many years in bovine practice for diagnosing inflammatory diseases. GT is used primarily to demonstrate increased serum concentrations of fibrinogen and globulin. Glutaraldehyde binds with free amino groups in fibrinogen and immunoglobulin to create a clot in a first degree chemical reaction. The clotting time of the GT estimates the content of proteins produced in response to inflammation. The applicability of GT for diagnosing inflammation in the horse has never been investigated. The objective of t...
Cissell JM, Milton SC, Dahlgren LA.To evaluate the effects of prostaglandin E₂ (PGE₂) treatment on the metabolism of equine tendon fibroblasts in vitro to aid in investigating the response of tendon fibroblasts to injury and novel therapeutics. Methods: Superficial digital flexor tendon fibroblasts isolated via collagenase digestion from six young adult horses were grown in monolayer in four concentrations of PGE₂ (0, 10, 50, 100 ng/ml) for 48 hours. Cells and medium were harvested for gene expression (collagen types I and III, cartilage oligomeric matrix protein [COMP], decorin, and matrix metalloproteinase-1, -3, and -1...
Langton CM, Riggs CM, Evans GP.The velocity of ultrasound waves through bone has been used widely as a non-invasive method for assessing bone quality. Accurate measurement of velocity depends on accurate assessment of the distance travelled by the sound wave. It has been argued that the sonic pathway is deflected around the marrow cavity and so does not follow a straight line through the bone; therefore, correction factors have been developed to account for the extra distance travelled. This hypothesis was examined in vitro using sections from the equine third metacarpal bone. Two 1 MHz transducers used with the transmittin...
Fenwick BW, Schore CE, Osburn BI.Equine, caprine, ovine, canine and feline peripheral blood lymphocytes were evaluated in a short term dose-response study for their in vitro blastogenic responsiveness to human recombinant interleukin-2(125) (HrIL-2(125] alone or in combination with phytohemagglutinin-P, concanavalin-A, and pokeweed mitogen. HrIL-2(125) induced lymphocyte proliferation in all of the animals tested. The magnitude of the proliferative response varied among the species of animal tested. In all cases the proliferative response was dependent on the concentration of HrIL-2(125). HrIL-2(125) at a minimum concentratio...
Miller SK.1. Results of laboratory experiments which compared horse and human serum butyrylcholinesterase (EC 3.1.1.8) with respect to their acid inactivation and ammonium sulfate protection show: 2. Horse serum butyrylcholinesterase is more resistant to inactivation at pH 3.0 than human serum butyrylcholinesterase. 3. The loss of activity at pH 3.0 for both horse and human butyrylcholinesterase does not follow first order kinetics. 4. Both human and horse serum butyrylcholinesterase are protected from pH 3.0 inactivation by ammonium sulfate concentrations up to 33% saturation (1.37 M).
Figueiredo MD, Salter CE, Hurley DJ, Moore JN.Lipopolysaccharide-binding protein (LBP) is an acute phase protein that binds the lipid A moiety of lipopolysaccharide (LPS) and transfers LPS monomers to soluble CD14 in plasma or membrane bound CD14 on mononuclear phagocytes. The result of these interactions is activation of the TLR4 receptor complex, and the synthesis and release of inflammatory mediators. Inclusion of LBP in cellular assays increases the sensitivity of cells expressing CD14 to LPS. Therefore, the objectives of this study were to (1) compare differentially treated sera from cattle and horses as sources of LBP activity using...
Duthie S, Mills H, Burr P.Infection with equine arteritis virus is a notifiable disease with sporadic occurrence in the UK. As stallions may harbour the virus after infection, horses are screened for exposure by serological testing prior to breeding. The virus neutralisation test is considered the 'gold standard' serological screening test, but it is time-consuming and labour intensive; consequently there is a move towards more rapid screening methodology. In this study, a commercially available EVA antibody ELISA is assessed. The ELISA performed poorly with a specificity [corrected] of 26% and a sensitivity [corrected...
Yamamoto M, Nakagawa K, Fujiwara K, Shimizu A, Ikeguchi M, Ikeguchi M.Equine β-lactoglobulin (ELG) assumes non-native helices during refolding and in partially folded states. Previously, circular dichroism (CD) combined with site-directed mutagenesis identified helical regions in the acid- and cold-denatured states of ELG. It is also known that a fragment of ELG, CHIBL (residues 88-142), has a structure similar to that of the cold-denatured state. For the study reported herein, the structure of a shorter fragment, CHIBLΔF (residues 97-142), was investigated by CD and nuclear magnetic resonance spectroscopy. The secondary chemical shifts clearly showed that non...
Ellerbrock R, Canisso I, Feijo L, Lima F, Shipley C, Kline K.Urospermia is known to affect semen quality in many mammals, including stallions. Determinations of semen pH and creatinine and urea concentrations have been used to diagnose urine contamination in raw stallion semen. Unfortunately, practitioners suspecting urine contamination in cooled-shipped samples have no proven means to confirm the presence of urine. Therefore, the objectives of this study were (1) to assess the effects of urine contamination on sperm motility of extended fresh and cooled-stored stallion semen, (2) to evaluate the usefulness of semen color, odor, pH, and creatinine and u...
Moslemi S, Dintinger T, Dehennin L, Silberzahn P, Gaillard JL.The ability of human and equine placental microsomes to aromatize 7 alpha-methyl-19-nortestosterone (MNT) was studied. Kinetic analysis indicates that MNT shares the androgen-binding site of human and equine placental microsomal aromatases. Human placental microsomal estrogen synthetase had about a 2.5-fold higher relative affinity for MNT than the equine placental enzyme (KiMNT/Km androstenedione of 32 versus 87). However, MNT was not metabolized by human placental microsomes, whereas it was very actively metabolized by equine placental microsomes. Further studies using purified equine cytoch...
