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.
Novelli EL, Rodrigues NL, Chiacchio SB.Biochemical values are widely related with environmental agents, sex and age, and are used in disease diagnosis. Numerous reports have been published on the biochemical parameters of different breeds of horses. However, there is a paucity of information concerning Cu-Zn superoxide dismutase (SOD), ceruloplasmin, copper and zinc determinations in the serum. Blood samples from a total of 60 horses of the Mangalarga-Paulista breed, representing three age groups (0 to 4 months old, 6 to 18 months old and adult) were examined. Male horses have a higher mean value of SOD, ceruloplasmin and copper th...
Couture L, Lemonnier JP, Troalen F, Roser JF, Bousfield GR, Bellet D, Bidart JM.The equine (e) placental glycoprotein hormone eCG plays a critical though not completely understood role during the first trimester of gestation in mares. In the present work, we have developed immunoradiometric assays (m-IRMAs) for detection of eCG, eCG alpha, and eCG beta using combinations of monoclonal antibodies (mAbs) specific for epitopes that reside on free and/or combined subunits. The free eCG alpha m-IRMA was based on AHT20 mAb, specific for the free alpha-subunit of all species, and 125I-labeled ECG01 mAb, which recognizes both free and combined alpha-subunit from equine and primat...
Corstvet RE, Gaunt SD, Karns PA, Burgermeister D, McBride JW, Nicholson SM, Battistini RA.Four horses were inoculated with Ehrlichia risticii contained in either infected murine P388 D1 cells or heparinized blood from an infected horse. All 4 horses produced serum antibody, plasma antigen, and clinical signs of the disease. An enzyme-linked immunosorbent assay was used to detect antibody in the serum and was also used in conjunction with an anti-E. risticii monoclonal antibody to detect antigenemia. These laboratory and clinical findings were correlated to determine the efficiency of the antigen detection method for discerning E. risticii infection.
Rojas G, Vargas M, Robles A, Gutiérrez JM.Twenty batches of polyvalent antivenom produced at the Instituto Clodomiro Picado were analyzed for turbidity, both before and after freezing-thawing and lyophilization. Eight batches became turbid upon freezing-thawing, and this change correlated with high levels of cholesterol, triglycerides and lipoproteins, especially beta-lipoprotein. Since normal horse serum does not become turbid after freezing-thawing, despite the fact that it has high lipoprotein levels, the possibility was raised that phenol, used as a preservative during serum fractionation, might affect lipoproteins, inducing the a...
Reisen WK, Meyer RP, Presser SB, Hardy JL.The extrinsic incubation rate (inverse of the time in days from infection to median transmission) of western equine encephalomyelitis (WEE) and St. Louis encephalitis (SLE) viruses by laboratory strains of Culex tarsalis Coquillett increased as a linear function of incubation temperatures from 10 to 30 degrees C. The estimated temperatures for zero transmission thresholds (intercept of the X axis) were 10.9 and 14.9 degrees C, and the number of degree days above these thresholds required for median transmission (inverse of the slope) was 67.6 and 115.2, respectively. Although the bodies of mos...
Stitz L, Bilzer T, Richt JA, Rott R.Borna disease represents a unique model of a virus-induced immunological disease of the brain. Naturally occurring in horses and sheep, the mechanisms of pathogenesis have been studied in experimental animals, namely in the rat. Many investigations have revealed that the infection of the natural hosts principally follows the same pathogenic pathways as observed in rats, leading to a severe encephalomyelitis. This affliction of the central nervous system results in severe neurological disorders that again, are fully comparable in laboratory animals to those in the natural and the different expe...
Prosser CG, McLaren RD.Ligand blotting analysis of serum from the horse using radiolabelled IGF-I revealed a protein at 96 kDa which was not present in serum from goat, cow, sheep, deer or donkey. These latter species all displayed five labelled bands in the range 24 to 41 kDa. Conversely, these were only weakly labelled in serum from the horse. Size exclusion chromatography of horse serum pre-incubated with radiolabelled IGF-I revealed reduced binding in the 130-kDa peak compared with goat plasma, and ligand blotting analysis indicated the 96-kDa protein was present in this peak. The 96-kDa protein from horse serum...
Home WA, Ford JE, Gibson DM.Analysis of 10 cDNA encoding lambda L chains of horse Ig indicated that this species may employ a relatively small number of variable region (V lambda) genes in the splenic B cell population. The V lambda sequences of all of the cDNA analyzed were closely related (> 88% identity at the nucleotide level) and were characterized by a deletion of the amino acid residue at position 3 compared with V lambda sequences so far described in other species. The 10 V lambda sequences could be grouped into three groups, V lambda 1 to V lambda 3, on the basis of a number of linked substitutions. Sequences...
Orsini JA, Ramberg CF, Benson CE, Dreyfuss DJ, Vecchione JA, Kunz CC.Vancomycin hydrochloride was infused intravenously (i.v.) over a 30-min period in five horses at doses of 6.6, 11.0 and 15.4 mg/kg. Vancomycin concentration in plasma and synovial fluid samples was measured using a polarization immunoassay. A pharmacokinetic model was developed to accommodate the special features of the present study. The data were described by a two compartment open model with synovial fluid as an additional compartment in exchange with plasma. Minimum inhibitory concentration (MIC) and minimum bacterial concentration (MBC) were measured for Staphylococcus aureus and Enteroco...
Science (New York, N.Y.)December 11, 1992
Volume 258, Issue 5089 1748-1755 doi: 10.1126/science.1334573
Pelletier H, Kraut J.The crystal structure of a 1:1 complex between yeast cytochrome c peroxidase and yeast iso-1-cytochrome c was determined at 2.3 A resolution. This structure reveals a possible electron transfer pathway unlike any previously proposed for this extensively studied redox pair. The shortest straight line between the two hemes closely follows the peroxidase backbone chain of residues Ala194, Ala193, Gly192, and finally Trp191, the indole ring of which is perpendicular to, and in van der Waals contact with, the peroxidase heme. The crystal structure at 2.8 A of a complex between yeast cytochrome c pe...
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...
Vickers PJ, O'Neill GP, Mancini JA, Charleson S, Abramovitz M.To identify regions of 5-lipoxygenase-activating protein (FLAP) important for the function of the protein and the binding of leukotriene biosynthesis inhibitors, we performed a cross-species analysis of FLAP. FLAP from all 10 mammalian species analyzed (human, monkey, horse, pig, cow, sheep, rabbit, dog, rat, and mouse) were immunologically cross-reactive and specifically bound leukotriene biosynthesis inhibitors with high affinity. Using the polymerase chain reaction, cDNA clones for FLAP from six species (monkey, horse, pig, sheep, rabbit, and mouse) were isolated and sequenced. The deduced ...
Stowe HD, Herdt TH.Assessment of the selenium status of livestock is an important aspect of production medicine, but variations in reported values between laboratories and between methods may be > 30%. Reliable interpretations require considerable experience with an assay and an extensive database from field and research case samples of a variety of species. The Michigan State University Animal Health Diagnostic Laboratory (MSU-ADHL) has offered Se analyses by acid-digestion and fluorometric detection since 1982. This laboratory expects serum Se values (nanograms per milliliter) of livestock to increase graduall...
Craig AM, Blythe LL, Rowe KE, Lassen ED, Barrington R, Walker KC.Recent evidence concerning the pathogenesis of equine degenerative myeloencephalopathy indicated that low blood alpha-tocopherol values are a factor in the disease process. Variables that could be introduced by a veterinarian procuring, transporting, or storing samples were evaluated for effects on alpha-tocopherol concentration in equine blood. These variables included temperature; light; exposure to the rubber stopper of the evacuated blood collection tube; hemolysis; duration of freezing time, with and without nitrogen blanketing; and repeated freeze/thaw cycles. It was found that hemolysis...
Leong LM, Tan BH, Ho KK.Nonheme iron proteins can be visualized as blue bands in native polyacrylamide gels using a staining method that is both simple and rapid. The reaction of potassium ferricyanide with protein-bound iron atoms to form royal blue complexes occurs almost instantaneously and is sensitive enough to detect 1 microgram of analytical-grade ferritin and 2 micrograms of purified ferredoxin from cyanobacteria. No special treatment of reagents or apparatus was necessary. On comparison, this stain was found to be more specific than the Ferene S stain, not detecting bovine serum albumin even when present as ...
Thomas LA, Brown SA.The relationship between colloid osmotic pressure (COP) and protein concentration was investigated for purified proteins and plasma samples obtained from cattle, horses, dogs, and cats. At equivalent concentrations, bovine albumin exerted a COP that exceeded that of gamma-globulins by a mean factor of 4.4. Similar relationships between COP and protein were observed in the other species. Consequently, for a given total protein concentration, COP was dependent on the albumin/gamma-globulins ratio. A commonly used nomogram for estimating COP from protein concentration, the Landis-Pappenheimer equ...
Su X, Morris DD, Crowe NA, Moore JN, Fischer KJ, McGraw RA.We describe the production and purification of recombinant equine tumor necrosis factor alpha (rETNF alpha), generation and characterization of murine monoclonal antibodies (Mabs) and rabbit polyclonal antibodies (Pabs) against ETNF alpha, and development of a sensitive enzyme-linked immunosorbent assay (ELISA). Genomic-derived DNA sequences encoding mature ETNF alpha were reconstructed by the polymerase chain reaction (PCR) and oligonucleotide-directed mutagenesis and were cloned into the vector pFLAG-1 for expression in Escherichia coli. rETNF alpha was purified by anti-FLAG immunoaffinity c...
Bousfield GR, Ward DN.Ovine (o) and equine (e) LH alpha-subunits were reduced and reoxidized using conditions known to be effective for bovine and human alpha-subunits. The major product of oLH alpha refolding was alpha-subunit monomer. In contrast, eLH alpha formed a 121,000 mol wt aggregate. Monomeric eLH alpha was recovered, but in greatly reduced yield. To test the effects of carbohydrate variation on the aggregation of equine alpha-subunits, all of the equine gonadotropin alpha-subunits (eFSH alpha, eCG alpha, eLH alpha, and free alpha-subunit) were reduced and reoxidized. In each case, the major product was t...
Le Goff D, Hannan J, Maboundou JC, Ayrault-Jarrier M.Equine plasma lipoproteins were fractionated into VLDL, LDL-1, LDL-2 and HDL by density gradient ultracentrifugation. From each lipoprotein fraction, five apo C like peptides of approx. M(r) 1400, 10000, 9500, 9000 and 8000 were detected by SDS-polyacrylamide gel electrophoresis. After partial purification by Sephadex G-75, one fraction, showing a strong activation of lipoprotein lipase, was further purified by Mono Q anion exchange column. Two of the apo C like peptides (M(r) 10000 and 8000) activated the bovine milk lipoprotein lipase in vitro; only one (M(r) 9500) inhibited the lipolytic ac...
Malark JA, Peyton LC, Galvin MJ.Peritoneal fluid and blood was collected from 8 healthy adult horses. Four 1-ml aliquots of peritoneal fluid from each horse were then contaminated with 0 ml (normal), 0.05 ml (1 drop), 0.10 ml (2 drops), and 0.20 ml (4 drops) of blood from the same horse. Samples were analyzed for RBC count, nucleated blood cell count, total protein concentration, and nucleated cell differential count. Statistical analysis revealed no significant changes in nucleated cell number, nucleated cell differential, or total protein concentration in peritoneal samples contaminated with blood. The RBC count significan...
Seahorn TL, Cornick JL, Cohen ND.The medical records of 75 horses with duodenitis-proximal jejunitis (DPJ) were reviewed. Ages, physical parameters, laboratory values, and treatment data were compared between horses surviving DPJ and horses not surviving DPJ (Table 1). Fifty of 75 horses (66.6%) survived. Sixty-six horses (88.0%) were managed with medical treatment alone and nine horses (12.0%) were managed with medical treatment plus surgical intervention. Using a logistic regression model, the association of each of the 19 physical and laboratory parameters with death was evaluated retrospectively in the 75 horses. Three pa...
Miller CC, Fayrer-Hosken RA, Timmons TM, Lee VH, Caudle AB, Dunbar BS.This study was designed to explore the composition of the equine zona pellucida (EZP) by one- and two-dimensional polyacrylamide gel electrophoresis (1D- and 2D-PAGE), silver staining and immunoblotting techniques. Antral follicles palpable on frozen-thawed equine ovaries were aspirated with a needle and syringe, and the resultant follicular fluid, cellular material and oocytes were pooled. Oocytes were placed in Petri dishes, moved by narrow-bore pipette to droplets of phosphate-buffered saline (PBS) and mechanically cleaned of cumulus cells. The EZP from these collected oocytes was solubiliz...
Mutlib AE, Chui YC, Young LM, Abbott FS.The metabolic fate of xylazine, 2-(2,6-dimethylphenylamino)-5,6-dihydro-4H-1,3-thiazine, in horses is described. The major metabolites identified in the hydrolyzed horse urine were 2-(4'-hydroxy-2',6'-dimethylphenylamino)-5,6-dihydro-4H-1,3-thiazi ne, 2-(3'-hydroxy-2',6'-dimethylphenylamino)-5,6-dihydro-4H-1,3-thiazi ne, N-(2,6-dimethylphenyl)thiourea, and 2-(2',6'-dimethylphenylamino)-4-oxo-5,6-dihydro-1,3-thiazine. These metabolites were also produced by incubating xylazine with rat liver microsomes. The major metabolite produced in vitro by rat liver preparations was found to be the ring op...
Eberhardt C, Gerhards H.The three tests (EQUI Z-Test, AGLUTINADE FOAL IMMUNITY, CITE Foal IgG-Test) were evaluated for their accuracy and usefulness in the field. Single radial immunodiffusion was used as reference method. All tests were easily and rapid to perform and results were obtained within a few minutes. It was easy to get the results of the CITE Foal IgG-Test, but use of the EQUI Z-Test and the FOAL AGLUTINADE IMMUNITY-Test needed some practice to get correct results. Results obtained by the CITE Foal IgG-Test correlated to single radial immunodiffusion in 94%, those obtained by FOAL AGLUTINADE IMMUNITY-Test...
Mendoza L, Nicholson V, Prescott JF.Reactions to Pythium insidiosum by sera from horses with active pythiosis were investigated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting. Five strains of P. insidiosum were grown in nutrient broth and then sonicated. After centrifugation, supernatant antigens were separated by SDS-PAGE. An exoantigen of Conidiobolus coronatus was also tested. Bands with molecular weights between 97,000 and 14,000 were identified by Coomassie blue and silver staining. After being transferred to nitrocellulose, the antigens were reacted against sera from six horses w...
de Vries AA, Chirnside ED, Horzinek MC, Rottier PJ.We have recently shown that the genome of equine arteritis virus (EAV) contains seven open reading frames (ORFs). We now present data on the structural proteins of EAV and the assignment of their respective genes. Virions are composed of a 14-kDa nucleocapsid protein (N) and three membrane proteins designated M, GS, and GL. M is an unglycosylated protein of 16 kDa, and GS and GL are N-glycosylated proteins of 25 and 30 to 42 kDa, respectively. The broad size distribution of GL results from heterogeneous N-acetyllactosamine addition since it is susceptible to digestion by endo-beta-galactosidas...
Agius CT, Nagesha HS, Studdert MJ.A new equine herpesvirus, provisionally designated equine herpesvirus 5 (EHV5; Browning and Studdert (1987) J. Gen. Virol. 68, 1441-1447), was examined for the degree of genomic difference from equine herpesvirus 2 (EHV2) by Southern hybridizations. EHV5 and EHV2 whole genomic DNA probes were highly specific for homologous DNA only, indicating that significant genomic difference exists between the two viruses. Restriction endonuclease analysis of EHV5 strain 2-141 (EHV5.2-141) revealed that the genome is 179 kb and exists as a single isomer. Clones representing 82% of the genome were obtained ...
Ranz AI, Miguet JG, Anaya C, Venteo A, Cortés E, Vela C, Sanz A.A panel of 32 hybridoma cell lines secreting monoclonal antibodies (MAbs) reactive with African horsesickness virus serotype 4 (AHSV-4) has been developed. Four of the MAbs recognized the major core antigen VP7, twenty recognized the outer capsid protein VP2 and eight reacted with the non-structural protein NS1. With the VP7-specific MAbs a rapid and sensitive double antibody sandwich immunoassay has been developed to detect viral antigen in infected Vero cells and in spleen tissue from AHSV-infected horses. The sensitivity of the assay is 10 ng viral antigen per 100 microliters. The NS1-speci...
Bermúdez VM, Miller RB, Rosendal S, Fernando MA, Johnson WH, O'Brien PJ.The cytopathic effects induced by five strains of Mycoplasma equigenitalium for cells of equine uterine tube explants were tested by measuring changes in cellular and extracellular concentrations of calmodulin (CaM). Calmodulin concentrations in samples of total homogenate (TH) and total homogenate supernates (THS) of the infected equine uterine tube explants were significantly lower than respective measurements on noninfected controls. In tissue culture medium fractions (TCM) of some infected explants, CaM concentrations were significantly higher than noninfected controls (p > 0.95). The r...
Callan MB, Giger U, Oakley DA, Scotti MV, Shofer FS.In veterinary medicine, PCV determined by centrifugation of blood in a microhematocrit tube is the most common clinical test used to initially assess and monitor anemic and polycythemic animals. In contrast, blood hemoglobin (Hb) concentration, rather than PCV, is generally determined in human patients. One automated system photometrically measures blood Hb concentration after conversion of Hb to azide methemoglobin without dilution and was found to be a simple and accurate instrument for use in human medicine. We evaluated the system for its accuracy in measuring blood Hb concentration in ani...
Van de Water E, Oosterlinck M, Duchateau L, Pille F.The purpose of this study was to determine whether the scil Vet abc Plus(+) (SCIL Animal Care Company, Altorf, France), an impedance hematology analyzer, can accurately quantify and differentiate nucleated blood cells (NBCs) in equine synovial fluid. Synovial fluid samples (n=242) in different stages of experimentally induced inflammation were analyzed with and without hyaluronidase pretreatment and compared to manual hemocytometer counts and smear reviews. No significant effect of hyaluronidase pretreatment was observed. Total nucleated cell counts of the scil Vet abc Plus(+) were significant...
Hodgson DR, Rose RJ, Allen JR, Dimauro J.Muscle biopsy samples were collected from the left middle gluteal muscle of the horses participating in day 2 (speed and endurance test) of a three day event. Six Thoroughbred horses were biopsied the day before and within 30 minutes of completion of the speed and endurance test. Serial muscle sections were reacted histochemically for myosin adenosine triphosphatase activity after acid pre-incubation to demonstrate Type I, IIA and IIB fibers and the glycogen content in the individual fibers was assessed using the periodic acid Schiff (PAS) reaction. Total glycogen in muscle was measured fluoro...
Moeller BC, Sams RA, Guinjab-Cagmat J, Szabo NJ, Colahan P, Stanley SD.Testosterone is an anabolic androgenic steroid (AAS) that is endogenously produced by both male and female horses that also has the potential for abuse when administered exogenously to race horses. To recommend appropriate withdrawal guidelines so that veterinarians can discontinue therapeutic use prior to competition, the pharmacokinetics and elimination of testosterone were investigated. An aqueous testosterone suspension was administered intramuscularly in the neck of Thoroughbred horses (n = 20). The disposition of testosterone from this formulation was characterized by an initial, rapid a...
Roser JF, Chang YS, Papkoff H, Li CH.A specific and sensitive homologous radioimmunoassay has been developed for equine prolactin, suitable for measuring prolactin concentrations in serum of horses. The sensitivity of the assay ranged from 0.4 to 0.6 ng/ml and the intra- and inter-assay coefficients of variation averaged 6.9 and 15.4%, respectively, for five doses of hormone. Cross-reactivity with other mammalian and nonmammalian prolactins and growth hormones was less than 20 and 0.3%, respectively. Cross-reactivity with equine growth hormone was less than 0.07%. Equine serum and pituitary extracts showed parallel dilution-respo...
Maurel MC, Ban E, Bidart JM, Combarnous Y.In the present study we have established an immunochemical mapping of equine Chorionic Gonadotropin (eCG/PMSG) using three monoclonal antibodies (mAbs), namely the antibodies ECG01, E10 and D7, raised against the native hormone. These antibodies do not bind to reduced, alkylated hormone, suggesting that they recognize discontinuous rather than continuous epitopes. We have also assessed the reactivity of mAbs towards human CG, and ovine, porcine, equine and bovine LH and FSH. The antigenic determinant recognized by ECG01 is localized on the alpha-subunit of equine gonadotropins and of human CG ...
Ek N.Selected transferrin phenotypes from 14 horses were investigated by antigen-antibody crossed electrophoresis. Horse sera were subjected to starch gel electrophoresis followed by right angle electrophoresis in agarose gels containing rabbit produced anti-horse transferrin. This technique gave an additional zone in the front as compared with 2 transferrin zones seen after ordinary starch gel electrophoresis. Comparisons of transferrin concentrations in horse sera were performed by an immunodiffusion technique. Values were related to a chosen reference serum. A total of 372 horses (210 Norwegian ...
Choi YH, Chung YG, Seidel GE, Squires EL.The objective was to compare culture media for in vitro maturation of equine oocytes and for in vitro culture of zygotes produced from IVF of partially zona-removed oocytes. Cumulus-oocyte complexes from slaughterhouse-derived ovaries were washed in m-Dulbecco's PBS and cultured in TCM-199, F10-DMEM or c-F10-DMEM (50% F10-DMEM + 50% F10-DMEM conditioned medium from culture of an equine trophoblast monolayer for 3 or 4 days). All media included FSH, LH, E2, and 10% FCS. After 28 to 30 h maturation, cumulus expansion was scored from 0 (no expansion) to 4 (fully expanded). Oocytes with a 1st pola...
Wernery U, Rodriguez M, Raghavan R, Syriac G, Miriam Thomas M S, Elizabeth SK, Federico Ronchi G, Muhammed R, Patteril NA, Joseph S.African horse sickness (AHS) is a devastating viral disease of equids that was first recorded in 1327. Currently, prevention and control of the disease are based on attenuated vaccines and midge control. It has been shown that attenuated Orbivirus vaccines are not always safe as they may reverse to virulence. Objective: In the Emirate of Dubai, a vaccination experiment was carried out with an inactivated AHS vaccine produced at the Central Veterinary Research Laboratory (CVRL), Dubai, UAE to investigate the humoral antibody response of AHS-naïve horses to this vaccine. Our vaccination experim...
Platt D, Bayliss MT.The effect of human recombinant insulin-like growth factor 1 (rhIGF-1) on proteoglycan (PG) metabolism of full thickness equine articular cartilage explants was investigated. PG synthesis was stimulated at all ages, but higher concentrations of rhIGF-1 were required for maximal stimulation of adult cartilage. There were no changes in the hydrodynamic size, electrophoretic heterogeneity or composition of proteoglycans isolated from rhIGF-1-stimulated cartilage. rhIGF-1 reduced the rate of turnover of both newly synthesized and endogenous proteoglycans in all ages of cartilage investigated. The ...
Reyner CL, Winter RL, Maneval KL, Boone LH, Wooldridge AA.To investigate the effects of recombinant equine IL-1β on function of equine endothelial colony-forming cells (ECFCs) in vitro. Methods: ECFCs derived from peripheral blood samples of 3 healthy adult geldings. Methods: Function testing was performed to assess in vitro wound healing, tubule formation, cell adhesion, and uptake of 1,1'-dioctadecyl-3,3,3',3' tetramethylindocarbocyanine perchlorate-labeled acetylated low-density lipoprotein (DiI-Ac-LDL) by cultured ECFCs. Cell proliferation was determined by 2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide assay. Effects o...
McKinney AR, Suann CJ, Dunstan AJ, Mulley SL, Ridley DD, Stenhouse AM.The equine phase I and phase II metabolism of the synthetic anabolic steroid stanozolol was investigated following its administration by intramuscular injection to a thoroughbred gelding. The major phase I biotransformations were hydroxylation at C16 and one other site, while phase II metabolism in the form of sulfate and beta-glucuronide conjugation was extensive. An analytical procedure was developed for the detection of stanozolol and its metabolites in equine urine using solid phase extraction, acid solvolysis of phase II conjugates and analysis by positive ion electrospray ionization ion ...
Runcan EE, Pozor MA, Zambrano GL, Benson S, Macpherson ML.The acrosome is a highly specialised region of the spermatozoon that is essential for fertilisation. Defects or dysfunction of this structure have been associated with fertility problems in man and various domestic species including stallions. Current methods of evaluating the acrosome of stallion spermatozoa are time consuming and require specialised equipment, which is cost prohibitive to the average practitioner. Objective: To evaluate 2 conventional stains (Dip Quick and Spermac) and determine their usefulness in assessing acrosome integrity in stallions as compared with specific acrosomal...
Stewart AJ, Hardy J, Kohn CW, Toribio RE, Hinchcliff KW, Silver B.To evaluate the diagnostic value of serum concentrations of total magnesium (tMg) and ionized magnesium (iMg), concentrations of magnesium (Mg) in muscle, intracellular Mg (icMg) concentrations, urinary Mg excretion (EMg), Mg clearance (CMg), and fractional clearance of Mg (FCMg) in horses fed diets with Mg content above and below National Research Council recommendations. Methods: 9 young female horses. Methods: 6 horses were fed a reduced-Mg diet for 29 days followed by an Mg-supplemented diet for 24 days. Control horses (n = 3) were fed grass hay exclusively. Blood, urine, and tissue sample...
Jain NC, Vegad JL, Kono CS.Equine neutrophil antibody was raised in rabbits inoculated with equine neutrophils isolated to purity greater than 99.0%, using Percoll density-gradient sedimentation. Neutrophil antibody was detected by use of agar gel diffusion, leukoagglutination, indirect immunofluorescence, staphylococcal protein A and streptococcal protein G binding, and phagocytic inhibition techniques. Precipitin lines and leukoagglutination were seen in antiserum dilutions of 1:4 and 1:64, respectively. The specific nature of leukoagglutination was characterized by the formation of rosette-like clumps of neutrophils....
Coyne CP, Hornof WJ, Kelly AB, O'Brien TR, DeNardo SJ.Two methods were analyzed for the rapid extraction of equine fibrinogen from fresh plasma, using ammonium sulfate-sodium phosphate buffer. Fibrinogen from each of these 2 methods was then radiolabeled with 125I (half-life = 60.2 days, gamma = 35 keV), using monochloroiodine reagent. Mean protein-bound activity was 98.5% and mean clottable radioactivity was 94.1%. Radiolabeled fibrinogen administered IV to 15 horses had an overall mean (+/- SD) plasma half-life of 4.95 +/- 0.44 days.
Horney BS, Honor DJ, MacKenzie A, Burton S.Serum sorbitol dehydrogenase (SDH) activities in 10 cows and nine horses were measured using an automated clinical analyzer. The serum samples were divided into aliquots that were stored at room temperature (21 degrees C), refrigerated (0-5 degrees C), or frozen (-30 degrees C). The stability of the SDH activity was monitored at various intervals. SDH activity in bovine sera remained stable for at least 5 hours at room temperature, 24 hours refrigerated, and 72 hours frozen without any significant (p < 0.05) differences from the initial serum values. In equine sera, SDH activity remained st...
Lopes-Pereira CM, Harun M, Schmidtova D, Timane I, Mabasso M, Otto F, Bogin E.Adapting the concept of "bed-side" patient analysis, the Boehringer-Mannheim Reflotron was evaluated for its possible use in veterinary medicine under outdoor-field conditions. Horse blood was analysed with the Cobas Bio analyzer, and indoor and outdoor analyses were also performed with the Reflotron. All values showed close agreement with no significant differences. Good correlation coefficients (r values around 0.9000) were also seen between all methods used. The Reflotron was operated under outdoor-field conditions by using, whenever available, an on-farm electricity source, or a gas operat...
Baeyens WR, Van der Weken G, Haustraete J, Aboul-Enein HY, Corveleyn S, Remon JP, García-Campaña AM, Deprez P.The group of LiChrospher ADS (alkyl-diol silica) sorbents that make part of a unique family of restricted-access materials, have been developed as special packings for precolumns used in the LC-integrated sample processing of biofluids. The advantage of these sorbents lies in the direct injection of untreated biological fluids, that is without sample clean-up, the elimination of the protein matrix with a quantitative recovery together with an on-column enrichment. The present method is based on previous work applying UV detection at 260 nm for ketoprofen determinations. Plasma samples introduc...
Mills JS, Kinsley MA, Peters DF, Weber PSD, Shearer TR, Pease AP.The purpose of this study was to determine whether there was a correlation between circulating and intra-synovial Dkk-1 and radiographic signs of equine osteoarthritis. Methods: Circulating and intra-synovial Dkk-1 levels were measured in clinical cases using a commercially available human Dkk-1 ELISA. Radiographs were performed of the joints from which fluid was collected and these were assessed and scored by a boarded radiologist for joint narrowing, subchondral bone sclerosis, subchondral bone lysis, and periarticular modelling. Comparisons were made between radiographic scores and the conc...
Taylor P, Scarth JP, Hillyer LL.Within equine drug surveillance, there is significant interest in analyzing intact phase II conjugates of drugs in urine, but progress has been limited by a lack of reference material. Methods: In this study, in vitro techniques using equine liver fractions were employed to produce glucuronide and sulfate conjugates of stanozolol, 16β-hydroxystanozolol and nandrolone, the glucuronide conjugate of morphine and the glutathione metabolite of chlordinitrobenzene for the first time in equine sports drug surveillance. Results: The glucuronide conjugate of the synthetic progestagen altrenogest was a...
Ruiz Silva BE, Burtnick LD.Gelsolin can be purified from horse blood plasma by treating the plasma sequentially with an anion-exchange medium in the presence and then the absence of free Ca2+. The purified gelsolin migrates as a 90-kilodalton protein on electrophoresis in polyacrylamide gels in the presence of sodium dodecyl sulfate. It has an absorption coefficient of 1.4 mL/(mg.cm) and is similar in amino acid composition to other plasma gelsolins. Horse plasma gelsolin has an intrinsic sedimentation coefficient of 4.8S and a Stokes' radius of 3.8 nm. Hydrodynamic calculations suggest it to be a rather globular protei...
Haywood PE, Teale P, Moss MS.Thoroughbred geldings were fed racehorse cubes containing a predetermined concentration of theobromine in the form of cocoa husk. They were offered 7 kg of cubes per day, divided between morning and evening feed, and food consumption was monitored. Urinary concentrations of theobromine were determined following the consumption of cubes containing 11.5, 6.6, 2.0 and 1.2 mg per kg of theobromine, to verify whether or not such concentrations would produce positive urine tests. Pre-dose urine samples were collected to verify the absence of theobromine before each experiment. It became apparent fro...
Weers JG, Maki AH.Triplet-singlet energy transfer has been studied in the complex formed between auramine O (AO) and horse liver alcohol dehydrogenase with optically detected magnetic resonance (ODMR) spectroscopy. The results show that Trp-15 and Tyr residues transfer triplet energy mainly by a trivial process, whereas Trp-314 transfers triplet energy by a Förster process with two observed lifetimes at 77 K of 170 and 50 ms. The different Förster energy-transfer lifetimes are ascribed either to quenching of the two Trp-314 residues of the dimer by a single asymmetrically bound AO or to two distinct conformat...
Noël PG, Couëtil L, Constable PD.Plastic heparinised vacutainer tubes are used for blood gas analysis in horses. This collection method may not be ideal because influx of atmospheric O(2) through the permeable plastic wall of the vacutainer tube and loss of CO(2) into the gas phase above the blood sample should increase blood PO(2) and decrease PCO(2), respectively. Objective: To determine the effects of collecting blood into plastic vacutainer tubes and storage conditions on blood gas analysis values. Methods: Blood was obtained from 6 healthy horses and tonometered at 37 °C with 12% O(2) and 5% CO(2). Three ml aliquots of ...
Reimer G, Suarez A, Chui YC.A standardbred mare was dosed with 40 mg yohimbine intravenously. Serum and urine samples were collected and analyzed for yohimbine using solvent extraction and reversed-phase high-performance liquid chromatography (HPLC) with fluorescence detection. Maximum yohimbine concentrations of 45 and 18 ng/mL were observed in serum and urine samples, respectively. Elimination was rapid, with half-lives of approximately 20 and 53 min observed for serum and urine, respectively. The presence of yohimbine in these samples was confirmed by liquid chromatography/mass spectroscopy (LC/MS/MS).
Desmadril M, Mitraki A, Betton JM, Yon JM.The unfolding-folding transition of phosphoglycerate kinase induced by GuHC1 was studied at equilibrium. Various signals were used to follow the transition: fluorescence emission, difference spectra, circular dichroism and enzymatic activity. The non-coincidence of transition curves obtained from different structural parameters indicate a deviation from a two-state process. The view that structural domains behave as independent "folding units" is critically discussed.
O'Niell FD, Issel CJ.Growth kinetics of equine influenza virus-A1, equine herpesvirus-1, and equine rhinovirus-1 were determined in susceptible cell monolayers and in organ cultures of equine fetal tracheal and nasal turbinate epithelium. Equine influenza virus-A1 was replicated in cell and organ cultures and was released more readily and for longer periods from nasal turbinate epithelium than from tracheal epithelium. Equine herpesvirus-1 was also replicated in cell and organ cultures. During the first 24 hours after inoculation, equine herpesvirus-1 was released more readily from tracheal epithelium than from na...
Samuel CA, Ricketts SW, Rossdale PD, Steven DH, Thurley KW.Endometrial biopsies obtained from mares at different stages of the oestrous cycle, during anoestrus and in various abnormal conditions were examined with the scanning electron microscope. Preliminary observations suggest that the patterns of secretory and ciliary activity in the uterine epithelium are similar to those observed by electron microscopical techniques in laboratory and other large domestic animals. The response of the epithelial cells to hormonal variations and infections is compared with that of the endometrium as seen with the light microscope.
Chen CL, Goldberg J, Gronwall RR.A radioimmunoassay was developed for prednisolone using IgG purified from rabbit antiserum. The assay was employed to determine the pharmacokinetics of prednisolone following intravenous administration of 450 mg of prednisolone sodium succinate (Solu Delta Cortef) to five adult Thoroughbred horses. The RIA had a sensitivity of 2 ng/ml and was relatively specific. It had cross-reactivity with 21-deoxycortisol (83.3%) cortisol (27.8%), 11-beta-hydroxyprogesterone (39.2%) and 17-hydroxyprogesterone (50%). However, it did not cross-react with naturally occurring steroids (cholesterol, testosterone...
Berhane Y, Bailey SR, Harris PA, Griffiths MJ, Elliott J.Elevated plasma homocysteine (HCy) concentration is a risk factor for cardiovascular diseases associated with endothelial dysfunction, including the human digital ischaemic disease, Raynaud's phenomenon. Objective: HCy causes dysfunction of equine vascular endothelium and elevated plasma concentrations predispose to laminitis. Objective: To determine 1) the concentration of HCy in vitro, which inhibits equine vascular endothelial cell function and 2) any association between risk of laminitis and plasma HCy concentration. Methods: Endothelial function was studied by measuring endothelium-depend...
