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.
Galosi CM, Norimine J, Echeverría MG, Oliva GA, Nosetto EO, Etcheverrigaray ME, Tohya Y, Mikami T.The genomes of 10 equine herpesvirus 1 (EHV-1) strains isolated in Argentina from 1979 to 1991, and a Japanese HH1 reference strain were compared by restriction endonuclease analysis. Two restriction enzymes, BamHI and BglII, were used and analysis of the electropherotypes did not show significant differences among isolates obtained from horses with different clinical signs. This suggests that the EHV-1 isolates studied, which circulated in Argentina for more than 10 years, belong to a single genotype.
Chua HC, Stewart B, Lim BH, Lee HK.A chromatographic method was developed to detect and confirm the presence of chlorpropamide (I) in horse plasma samples, for antidoping control. The plasma sample (1 ml) was extracted with dichloromethane and screened by high-performance liquid chromatography, and confirmation of the drug's presence was accomplished by using gas chromatography-mass spectrometry (GC-MS). The limit of detection was found to be 3.5 ng/ml at a signal-to-noise ratio of three. Derivatization of I with N,O-bis-(trimethylsilyl)trifluoroacetamide with 1% trimethylchlorosilane allowed for highly stable, accurate and sen...
Mikami T, Tsuchihashi K, Kashiwagi M, Yachida Y, Daino T, Hashi K, Akino T, Gasa S.A sulfatide, O-fatty-acylated 3-sulfogalactosylceramide at C6-O on galactoside, was isolated from equine brain and the chemical structure was characterized by proton NMR and MS. The O-acylation site of the acylated sulfatide was determined by the down-field shift of protons attached to a carbon having an O-acyl group in the NMR spectrum and by analysis of a partially methylated derivative before and after acetalization of the intact sulfatide using GC-MS. The O-acyl chain length was determined by GLC, revealing that it exclusively had palmitoyl and stearoyl residues as the major fatty acids. T...
Mobarak MS, Ryan MF.Adult Strongylus vulgaris, collected from the caecum of infected horses and embedded in paraplast using standard methods, were sectioned for immunohistochemistry (IHC) studies. Antibodies were raised in rabbit against the excretory-secretory product (ESP) and against two constituent protein bands (28-30 kDa). The use of sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE), enzyme-linked immunosorbent assay (ELISA) and immunoblotting indicated the immunogenicity of ESP and of the subunits (28-30 kDa). In ELISA, both rabbit hyperimmune sera recognized the ESP and (28-30 kDa) ban...
Scepek S, Coorssen JR, Lindau M.Using the patch-clamp technique, we studied the role of protein phosphorylation and dephosphorylation on the exocytotic fusion of secretory granules with the plasma membrane in horse eosinophils. Phorbol 12-myristate 13-acetate (PMA) had no effect on the amplitude and dynamics of degranulation, indicating that the formation of fusion pores is insensitive to activation of protein kinase C (PKC). Fusion pore expansion, however, was accelerated approximately 2-fold by PMA, and this effect was abolished by staurosporine. Elevating intracellular Ca2+ to 1.5 microM also resulted in a 2-fold accelera...
Laidler P, Cowan DA, Houghton E, Kicman AT, Marshall DE.Recognition by the legal authorities that growth hormones (GHs) may be abused to improve sporting performance and/or physique has led to the implementation of controls that make it an offence to produce, supply, possess or import and export GHs, with intent to supply, without the authority to do so. A method is described for the discriminatory analysis of human, equine, porcine and bovine GHs for forensic purposes. Peptide-mass mapping by matrix-assisted laser desorption/ionization (MALDI) time-of-flight (TOF) mass spectrometry following tryptic digestion gave sequence coverages of 97.4%, 93.7...
Godard S, Schibler L, Oustry A, Cribiu EP, Guérin G.A horse BAC library was constructed with about 40,000 clones and mean insert size of 110 kb representing a 1.5 genome equivalent coverage and a probability of finding a single sequence of 0.75. It was characterized by PCR screening of about 130 sequences of horse microsatellites and exonic gene sequences retrieved from databases. BACs containing 8 microsatellites and 12 genes were subsequently localized by fluorescent in situ hybridization (FISH) on chromosomes. Two linkage groups were newly assigned to chromosomes: LG2 to ECA3 and LG5 to ECA24, and five linkage groups were also oriented--LG3,...
Hinterhofer C, Stanek C, Binder K.The elastic (E-) modulus of hoof horn samples as a function of moisture content was determined from different segments of the equine hoof. 110 hoof horn specimens with different pigmentation taken from six adult warm-blooded horses with no obvious pathological changes within t he foot were used for the 177 tension and bending tests which were performed in accordance with ASTM D 5026, ASTM D 5023 and DIN 53.457. E-moduli were determined under physiological conditions with mean 761.8, SD +/- 295.4 N/mm for dorsal wall samples, 708 +/- 280.4 N/mm2 for lateral wall samples, 230 +/- 92.4 N/mm2 for ...
Blackall PJ, Christensen JP, Bisgaard M.The objective of this work was to examine the diversity within Australian isolates of Actinobacillus equuli and related organisms by the genotypic method of ribotyping. Methods: Ribotyping, performed using the enzyme HaeIII, was used to examine the diversity in 12 field isolates of A equuli (five being capable of fermenting L-arabinose), one field isolate of Pasteurella caballi and two unclassifiable field isolates. Isolates were obtained from Australian horses, except for three isolates of A equuli (one L-arabinose positive and two L-arabinose negative) which were obtained from horses and a p...
Hammond SA, Issel CJ, Montelaro RC.Equine immunological research is hindered by the lack of a simple yet reliable general protocol by which to assay CTL activity specific for viral or parasitic antigens. We present here the first comprehensive analysis of the parameters necessary to reliably culture equine T cells and to analyze the antigen specific cytolytic activity of T lymphocytes utilizing the equine infectious anemia virus (EIAV) infection of outbred ponies as a source for in vivo primed T lymphocytes. Effective long-term in vitro culture of equine T cells was determined to require minimally 200 U/ml of recombinant human ...
Goudet G, Belin F, Bézard J, Gérard N.In the equine species, a large proportion of oocytes fail to complete meiosis during in-vitro culture. The biochemical and molecular basis of this failure is unknown. The meiotic cell cycle is controlled in part by the maturation-promoting factor (MPF) and the mitogen-activated protein kinase (MAPK). In this study, we evaluated the oocyte competence for in-vitro maturation and the expression of MPF components (p34cdc2 and cyclin B) and MAPK after in-vitro culture. The maturation rate was influenced by the culture medium and the physiological stage of the mare at the time of oocyte recovery. We...
Van den Hoogen BM, van de Lest CH, van Weeren PR, Lafeber FP, Lopes-Cardozo M, van Golde LM, Barneveld A.The purpose of this study was to determine whether changes in the synovial fluid (SF) induced by in vivo loading can induce an alteration in the metabolic activity of chondrocytes in vitro. Therefore, SF was collected from ponies after a period of box rest and after they had exercise for a week. Normal, unloaded articular cartilage explants were cultured in 20% solutions of these SFs for 4 days and chondrocyte activity was determined by glycosaminoglycan (GAG) turnover. In explants cultured in post-exercise SF, GAG synthesis was enhanced and GAG release was diminished when compared to cultures...
Constable PD, Hinchcliff KW, Muir WW.To compare the accuracy of anion gap (AG) and strong ion gap (SIG) for predicting unmeasured strong ion concentration in plasma and serum from horses. Methods: 6 well-trained Standardbred horses undergoing high-intensity exercise (experimental study) and 78 horses and ponies that underwent i.v. administration of lactic acid or endotoxin, and endurance, submaximal, or high-intensity exercise. Methods: Anion gap was calculated as AG = (Na+ + K+) - (Cl- + HCO3-), and SIG was calculated, using the simplified strong ion model, whereby SIG (mEq/L) = 2.24 x total protein (g/dl)/(1 + 10(6.65-pH)) - AG...
Lynch TM, Caron JP, Arnoczky SP, Lloyd JW, Stick JA, Render JA.To evaluate the influence of exogenous hyaluronan (HA) on in vitro synthesis of HA and collagenase by equine synoviocytes from normal and inflamed joints. Methods: 9 adult horses. Methods: Synoviocytes for culture were taken from the middle carpal joint of 3 horses with normal joints (control) and 6 horses with osteochondral fractures (principal). Synoviocytes were propagated in monolayer cultures and were incubated with 3 commercial HA products at concentrations of 0, 200, 400, and 1,500 micrograms/ml. Newly synthesized HA was radiolabeled with [3H]glucosamine and quantified by cetylpyridiniu...
Qin Z, Durand LG, Allard L, Cloutier G.In most studies that were aimed at evaluating the kinetics of red blood cell (RBC) aggregation, human blood was initially circulated at a high shear rate to disrupt the aggregates, and measurements were performed following a complete flow stoppage, during the process of rouleau formation. However, it is known that a very low shear rate can enhance the formation of aggregates, as demonstrated by the modal relationship of the shear-rate dependence of RBC aggregation. The objective of the present study was, thus, to evaluate the influence of sudden flow reductions compared to a complete flow stop...
Vidugiris GJ, Royer CA.The volume change for the transition from the native state of horse heart apomyoglobin to a pressure-induced intermediate with fluorescence properties similar to those of the well-established molten globule or I form was measured to be -70 ml/mol. Complete unfolding of the protein by pressure at pH 4.2 revealed an upper limit for the unfolding of the intermediate of -61 ml/mol. At 0.3 M guanidine hydrochloride, the entire transition from native to molten globule to unfolded state was observed in the available pressure range below 2.5 kbar. The volume change for the N-->I transition is relat...
Ebert DW, Roberts C, Farrar SK, Johnston WM, Litsky AS, Bertone AL.Measurements of absolute total reflectance were recorded from weight-bearing (n=9) and nonweight-bearing (n=9) equine articular cartilage specimens from 300 to 850 nm using a spectrophotometer with integrating sphere attachment. Following correction of measured spectra for interfacial reflections and edge losses, Kubelka-Munk theory was applied to estimate absorption and scattering coefficient, one-dimensional light intensity distribution, and light penetration depth. Kubelka-Munk absorption coefficients ranged from ∼7 cm-1 at 330 nm to ∼1 cm-1 at 850 nm. A localized absorption peak wa...
Takai S, Vigo G, Ikushima H, Higuchi T, Hagiwara S, Hashikura S, Sasaki Y, Tsubaki S, Anzai T, Kamada M.Polymerase chain reaction (PCR)-based assays were developed to detect virulent Rhodococcus equi in transtracheal aspirate samples from sick foals showing respiratory signs. An oligonucleotide primer pair from the sequence of the virulence-associated 15- to 17-kDa antigen gene of the virulence plasmid in virulent R. equi was used to amplify a 564 bp region by PCR, and the result was confirmed by Southern blot hybridization. No positive reaction was seen in DNA from 13 different microorganisms typically found in the respiratory tract. In tracheal aspirates seeded with virulent R. equi, a visible...
Ilobi CP, Nicolson C, Taylor J, Mumford JA, Wood JM, Robertson JS.Equine influenza viruses propagated in the laboratory in alternate hosts such as embryonated hens' eggs or mammalian cell culture have been analysed by HA sequencing and antigenically and their sequence compared to the original virus present in clinical material. In contrast to clinically derived human influenza virus which generally grows in MDCK cells without change, the data for equine influenza virus were less clear in that variants of equine virus were derived in both eggs and cells. The study indicated that the current use of eggs for equine influenza virus surveillance and vaccine produ...
Bird J, Larsen M, Nansen P, Kraglund HO, Grønvold J, Henriksen SA, Wolstrup J.Two sets of dung-derived organisms from soil routinely fertilized with manure (MA) and soil chemically fertilized (CH) were cultured separately in the laboratory. Baermannized organisms from these cultures were added to 20 g of faeces from strongyle-infected horses to form three treatment groups: (i) no soil organisms; (ii) low inoculum of soil organisms containing all organisms present in a suspension of approximately 100 adult female free-living nematodes; and (iii) high inoculum containing those soil organisms present with approximately 1000 adult female free-living nematodes. Three studies...
Peauroi JR, Fisher DJ, Mohr FC, Vivrette SL.A 14-year-old Arabian gelding had weight loss and anorexia of 3 weeks' duration. Results of repeated laboratory tests revealed persistent hypercalcemia and serum phosphorus concentration that was within or less than the reference range. Parathyroid hormone concentration was high. Histologic examination of specimens obtained at necropsy revealed parathyroid adenoma. A diagnosis of primary hyperparathyroidism attributable to a functional parathyroid adenoma was made. Abnormalities in calcium and phosphorus concentrations were similar to those seen with primary hyperparathyroidism in dogs, in whi...
Pellegrini A, Kalkinc M, Hermann M, Grünig B, Winder C, Von Fellenberg R.Equinins are a closely related group of proteins found in equine neutrophil granules. They demonstrate proteinase inhibiting activity restricted to microbial proteinase K and subtilisin, and they also possess antibacterial and antiviral properties. Antiproteinase K activity was measured in tracheobronchial secretions (TBS) of horses with mild (n = 15), moderate (n = 30) and severe (n = 16) chronic pulmonary disease, to determine its usefulness as an indicator of severity of disease and to measure neutrophil content. Determination of proteinase K inhibiting activity was based on a colorimetric ...
Brightwell G, Brown JM, Coates DM.Rt-PCR probes targeted to different gene sequences of VEE (Venezuelan equine encephalitis) virus strain TC-83 were assessed for their sensitivity, specificity and non-specific cross-reactivity. A generic VEE virus amplimer (VNSP4F2/VNSP4R2), targeted against nsP4 was identified, which was sensitive (detected at least 10 pfu) and robust (worked over a wide range of salt concentrations and annealing temperatures). An E2 amplimer designed against TC-83, (VE2F/VE2R), identified VEE strains TRD (1AB), P676 (1C), 3880 (1D) Everglades (2) vRNA whilst a second E2 primer pair designed against strain 68...
Sheoran AS, Lunn DP, Holmes MA.This paper describes the production of a panel of monoclonal antibodies (mAbs) identifying the four recognised equine IgG subisotypes IgG, IgGa, IgGb, IgGc and IgG(T). Pure preparations of the subisotypes for use in immunisations and testing were produced using a combination of gel filtration, salt precipitation, ion exchange chromatography and protein A and Protein G affinity chromatography. The specificity of mAbs for the IgG subisotypes was confirmed using ELISA assays, by characterisation of affinity purified proteins recognised by the mAbs, and by Western blotting of equine serum proteins...
Bhuyan AK, Udgaonkar JB.The unfolding kinetics of horse cytochrome c in the oxidized state has been studied at 10, 22, and 34 degreesC as a function of guanidine hydrochloride (GdnHCl) concentration. Rapid (millisecond) measurements of far-UV circular dichroism (CD) as well as fluorescence quenching due to tryptophan to heme excitation energy transfer have been used to monitor the unfolding process. At 10 degreesC, the decrease in far-UV CD signal that accompanies unfolding occurs in two phases. The unobservable burst phase is complete within 4 ms, while the slower phase occurs over tens to hundreds of milliseconds. ...
Raabe MR, Issel CJ, Montelaro RC.Equine infectious anemia virus (EIAV) has been shown to infect cells of monocyte/macrophage lineage. These primary cells are intrinsically difficult to obtain, to purify and to culture in vitro for extended periods of time. As a result, most in vitro studies concerning this lentivirus make use of primary equine fibroblasts or transformed canine or feline cell lines. We describe methods that yield reproducibly pure cultures of equine blood monocytes from peripheral blood mononuclear cells. The in vitro differentiation of these cells into mature equine macrophage was verified using various cytoc...
Waraksa E, Kowalski K, Kłodzińska E, Rola R, Ciekot J, Filipiak W, Bieńkowski T, Namieśnik J.Gamma-oryzanol (GO) has gained special attention in the equine sports industry in recent years due to its touted properties, including the fact that it may cause anabolic effects on muscle growth and reduce fatigue. Many manufactures offer supplements containing GO as a naturally occurring anabolic substance; however, some producers do not declare its presence in product compositions. Taking into consideration the touted properties of GO, its ambiguous effectiveness and the open character of the Prohibited Substances List established by the Fédération Equestre Internationale, there is an urg...
Rivero JL, Serrano AL, Henckel P.Biopsies of the gluteus medius muscle were taken at three different depths from 36 endurance horses aged 8.42 +/- 2.85 years and of both sexes. Twenty of the horses were considered to be excellent performers on the basis of the mean speed of their three fastest records in endurance events over the previous two or three years, whereas 16 were moderate performers. The biopsy samples were analysed for the activities of the enzymes citrate synthase (an indicator of citric acid cycle activity), 3-OH-acyl-CoA-dehydrogenase (an indicator of lipid oxidation) and lactate dehydrogenase (an indicator of ...
Hifumi T, Akioka K, Tanaka T, Miyoshi N.Alveolar echinococcosis, which is caused by a larval-stage infection of Echinococcus multilocularis, is a zoonosis with public health importance. Recently, alveolar echinococcosis in slaughtered horses has been reported in Japan and Poland. In terms of public health, a highly sensitive and specific diagnostic method is essential for early detection during meat inspection. In this study, the loop-mediated isothermal amplification (LAMP) assay was developed and validated to target the mitochondrial cytochrome b (cob) gene of E. multilocularis. Forty-one hepatic solid nodules obtained from each h...
Chang CK, Ward B, Ebina S.Carbon monoxide- and oxygen-binding rates and affinities were measured for horse heart myoglobins reconstituted with synthetic hemes lacking peripheral methyl and vinyl groups. There is an apparent correlation between heme size and ligand specificity, i.e. larger m values (ratios of CO vs O2 association rates, l'/k') with smaller hemes. However, this correlation broke down with the most dealkylated heme. This is interpreted as resulting from protein conformational changes altering the steric crowdedness at the O2-binding site. Spectral properties and autoxidation rates also corroborate this vi...
Cozzi B.A combined ultrastructural and immunocytochemical study was performed on the pineal gland of the horse in order to identify the cell types present and describe their characteristics. Comparisons have been made with other mammals. Two main cell types are present: pinealocytes and glial cells. Pinealocytes display different degrees of electron density in the nucleus and the cytoplasm, yet no ultrastructural feature supports the idea of separate populations. Putative secretory materials are stored in vesicles related to the Golgi apparatus. A variety of electron-dense bodies are present in the cy...
Ramos MI, Hermosura ME, Nakabayashi T.Horse, calf and bovine serum were successfully used as human serum substitutes in the in vitro cultivation of Plasmodium falciparum. Positive results were obtained only after gradually adapting the parasites to the substitute serum. Adapted lines were established within 4-5 weeks. 10% horse serum was observed to be the best substitute with growth rates comparable or even surprising that obtained in human serum. Pure calf or bovine serum supported stable growths of 20-30% less which was enhanced to comparable levels after addition of 1% glucose-peptone to the medium. Direct transfers of adapted...
Janda J, Lehmann M, Luttmann W, Marti E.Interleukin-3 is a growth and differentiation factor for various hematopoietic cells. IL-3 also enhances stimulus-dependent release of mediators and cytokine production by mature basophils. Function of IL-3 has not been studied in horses because of lack of horse-specific reagents. Our aim was to produce recombinant equine IL-3 and test its effect on sulfidoleukotriene and cytokine production by equine peripheral blood leukocytes (PBL). Equine IL-3 was cloned, expressed in E. coli and purified. PBL of 19 healthy and 20 insect bite hypersensitivity (IBH)-affected horses were stimulated with Culi...
Brown SA, Dusza K, Boehmer J.A relation exists between colloid osmotic pressure (pi) and serum total protein concentration; equations describing this relation have been used to determine a calculated value for pi. However, the relation between total protein concentration and pi is altered by the method used to measure protein and by changes in the ratio of concentrations of albumin (A) to globulin (G). We developed nomograms for estimating pi from A and G concentrations, using samples obtained from clinically normal animals and compared the accuracy of these nomograms with that of previously described equations relating p...
Khoshtariya DE, Dolidze TD, Shushanyan M, van Eldik R.Horse muscle myoglobin (Mb) was tightly immobilized at Au-deposited ~15-Å-thick mixed-type (1:1) alkanethiol SAMs, HS-(CH₂)₁₁-COOH/HS-(CH₂)₁₁-OH, and placed in contact with buffered H₂O or D₂O solutions. Fast-scan cyclic voltammetry (CV) and a Marcus-equation-based analysis were applied to determine unimolecular standard rate constants and reorganization free energies for electron transfer (ET), under variable-temperature (15-55 °C) and -pressure (0.01-150 MPa) conditions. The CV signal was surprisingly stable and reproducible even after multiple temperature and pressure cycl...
Viljanto M, Kaabia Z, Taylor P, Muir T, Habershon-Butcher J, Bailly-Chouriberry L, Scarth J.Boldenone is an anabolic-androgenic steroid that is prohibited in equine sports. However, in certain situations, it is endogenous or is believed to be formed by microbes in urine, and therefore, an approach for the differentiation is required. Following the identification of Δ1-progesterone and 20(S)-hydroxy-Δ1-progesterone as potential biomarkers of microbial activity, the presence of six steroids was investigated in the postrace urine of castrated male horses (geldings, n = 158). In line with endogenous findings from several other species when ultrasensitive methods are employed, bolde...
Taddei N, Buck M, Broadhurst RW, Stefani M, Ramponi G, Dobson CM.The stability and equilibrium unfolding behaviour of horse muscle acylphosphatase have been studied by denaturing the protein under various conditions of temperature, pH, and urea concentration. Far-ultraviolet circular dichroism (CD) and nuclear magnetic resonance (NMR) spectroscopy indicate that this small monomeric protein unfolds reversibly and cooperatively. Thermodynamic parameters, the Gibbs free energy delta G and enthalpy delta H of unfolding, have been estimated for denaturation of the protein from NMR and CD data as 19 kJ mol-1 and 350 kJ mol-1, respectively. CD and 1H-NMR results s...
Madry MM, Spycher BS, Kupper J, Fuerst A, Baumgartner MR, Kraemer T, Naegeli H.Compared to blood or urine, drugs can be detected for much longer periods in the long hair of horses. The aim of this study was to establish and validate a highly sensitive liquid chromatography tandem mass spectrometry (LC-MS/MS) method for the detection and quantification of frequently prescribed opioids, sedatives and non-steroidal anti-inflammatory agents in the mane and tail hair of horses. Based on an average growth rate of about 2 cm per month, times of administration reported by horse owners or veterinary physicians were related to drug localizations in hair. Hair samples were collecte...
Gao X, Jiang CG, Han XE, Zhao LP, Shen RX, Xiang WH, Zhou JH.To elucidate the function of the S2 gene in equine infectious anemia virus (EIAV) and its role in the attenuation of the Chinese attenuated EIAV vaccine strains, the S2 in the EIAV vaccine strain EIAV (FDDV) was reverse-mutated and the in vitro replication character of the resultant virus was evaluated. Based on the sequence variation of the S2 gene between the EIAV virulent strains and vaccine strains, all the four vaccine-specific sites in the S2 of an EIAV(FDDV) infectious clone, pFDDV3-8, were reverse-mutated to the sequences of the virulent strain EIAV(DV115). The reverse-mutated molecula...
Donné-Op den Kelder GM, de Haas GH, Egmond MR.At alkaline pH porcine pancreatic phospholipase A2 is known to bind two Ca2+ ions per protein molecule. One Ca2+ ion is strongly bound to the active site and is essential for enzyme activity. A second Ca2+ ion binds more weakly to the protein and improves the affinity of the enzyme for lipid-water interfaces severalfold at high pH values. A group having a pK around 6 controls enzyme binding to lipid-water interfaces in the absence of Ca2+. By use of proton titration techniques this group is now identified to be a carboxylate having an abnormally high pK. Its pK shifts to a value around 4.5 in ...
Heiskanen ML, Pirhonen A, Koskinen E, Mäenpää PH.The role of various environmental conditions on sperm motility and ATP content was investigated by incubating raw and washed spermatozoa collected with an open-ended artificial vagina from 10 stallions in various biological and artificial media under different atmospheric conditions. Spermatozoa did not survive for more than 12 h when kept unextended in the original seminal fluid in any circumstances. The most favourable media tested for long-term sperm survival were Kenney's medium or Kenney's medium supplemented with 10 mM-theophylline and 10 mM-Hepes, pH 7.2. Centrifugation and slow cooling...
Southwood LL, Gassert T, Lindborg S.It is the impression of some surgeons that geriatric horses have a lower survival rate compared to mature nongeriatric horses following colic surgery. One possible reason for this is that geriatric horses may be more critically ill at admission and have more severe disease than mature nongeriatric horses. Objective: To compare admission historical, physical examination and laboratory data for geriatric and mature nongeriatric horses referred for signs of colic. Methods: Medical records of horses admitted with a presenting complaint of colic between 2000 and 2006 were reviewed. Geriatric horses...
McDonald TL, Burger D.Initial separation and concentration of Clq from fresh, normal equine serum was accomplished by precipitation in 0.02 M acetate buffer, pH 5.5, at 4 degrees for 24 h. The re-dissolved precipitate was clarified by centrifugation at 80,000 g for 1 h and then dialysed against Tris-HCl buffer (0.05 M, pH 8.0) containing 10-3 M EDTA. The clarified dialysate remained biologically active at 5 degrees for at least 4 weeks. Biological activity of equine Clq was determined by assay of its ability to agglutinate sensitized sheep erythrocytes (EA). Following ammonium sulphate fractionation, Sepharose 4B g...
Salami F, Ghodrati M, Parham A, Mehrzad J.Bone marrow mesenchymal stem cells (BM-MSCs), as multipotent cells with self-renewal and plastic-adherent properties, have immunomodulatory effects on immune cells, including neutrophils. These cells are in close proximity in bone marrow (BM) sinusoids with non-multiplicative immature neutrophils. BM-MSCs exert their immunomodulatory effects on adjacent cells both directly (cell-to-cell contact) and indirectly (secretion of soluble factors). The aim of this study was to evaluate the effect of equine bone marrow mesenchymal stem cells (BM-MSCs) on the expression of some pro- and anti-apoptotic ...
Johnston JK, Morris DD.Sixty-eight horses with colic caused by small intestinal disease were allotted into 2 groups of 34 on the basis of recorded findings during exploratory celiotomy, necropsy, or response to medical treatment alone. Signalment, history, physical examination findings, and laboratory findings were compared between the group of horses with small intestinal obstruction and the group with duodenitis/proximal jejunitis. A significantly greater proportion of horses with duodenitis/proximal jejunitis were older than 2 years old (P less than 0.05). Differences in sex or breed distribution, or in seasonali...
Karakka Kal AK, Nalakath J, Kunhamu Karatt T, Perwad Z, Mathew B, Subhahar M.The detection and separation of medetomidine enantiomers from the complex biological matrices poses a great analytical challenge, especially in the field of forensic toxicology and pharmacology. Couple of researchers reported resolution of medetomidine using protein-based chiral columns, but the reported method is quiet challenging and tedious to be employed for routine analysis. This research paper reported a method that enables the enantio-separation of medetomidine by using polysaccharide cellulose chiral column. The use of chiralcel OJ-3R column was found to have the highest potential for ...
Deng L, Li Z, Tang C, Han Y, Zhang L, Liao Q.Equine pregnancy is currently diagnosed by rectal palpation, ultrasonographic examination, or by measuring changes in hormones in the blood. In the present study, we identified proteins that are differentially expressed in the sera of early pregnant and non-pregnant mares in order to develop a novel method for diagnosing equine pregnancy. Serum samples were obtained from 18 adult mares, pregnancy at day 32 after ovulation (n = 9) and in diestrus (n = 9). Proteomic analysis of the samples was conducted using liquid chromatography-electrospray ionization-tandem mass spectrometry. We iden...
Schnabel CL, Babasyan S, Freer H, Wagner B.Only few quantitative reports exist about the concentrations and induction of immunoglobulin A (IgA) in mucosal secretions of horses. Despite this, it is widely assumed that IgA is the predominant immunoglobulin on mucosal surfaces in the horse. Here, two new monoclonal antibodies (mAbs) against equine IgA, clones 84-1 and 161-1, were developed and characterized in detail. Both IgA mAbs specifically bound monomeric and dimeric equine IgA in different applications, such as Western blots and fluorescent bead-based assays. Cross-reactivity with other equine immunoglobulin isotypes was not observe...
Todhunter RJ, Wootton JA, Altman N, Lust G, Minor RR.Collagen type I and type II were purified from equine flexor tendon and articular cartilage, respectively. Equal amounts of these collagens were cleaved with cyanogen bromide, and 11 mixtures containing increasing proportions of type II collagen were separated in seven identical sodium dodecyl sulfate-polyacrylamide gels. The density of bands was measured in wet gels and the peak areas were used to form six ratios of peptide bands that had polynomial relationships with the known proportions of type I and type II collagen in the mixtures. Calibration curves for determining the proportion of typ...
Murray JA, Longland AC, Moore-Colyer MJ, Dunnett C.A series of experiments was conducted to determine the effects of a fibrolytic enzyme preparation (enzyme 1; E1) on the in vitro fermentation of lucerne incubated with equine faecal inocula. In experiment 1, high-temperature-dried (HT) lucerne was treated with five levels of E1 (0 to 2.4 ml/g DM) and incubated at 50 degrees C for 20 h. Samples then received a simulated foregut digestion (SFD) treatment before DM and NSP analysis. In experiment 2, HT lucerne was treated with the same enzyme levels used in experiment 1. Samples were then split into two groups; plus or minus an SFD treatment befo...
Dai X, Hilsen RE, Hu WG, Fulton RE.An electrochemiluminescence (ECL) immunoassay, incorporating chemically biotinylated and ruthenylated antibodies down-selected from a panel of monoclonal and polyclonal reagents, was developed to detect and identify Venezuelan equine encephalitis virus (VEEV). The limit of detection (LOD) of the optimized ECL assay was 10(3)pfu/ml VEEV TC-83 virus and 1 ng/ml recombinant (r) VEEV E2 protein. The LOD of the ECL assay was approximately one log unit lower than that of a sandwich enzyme-linked immunosorbent assay (ELISA) incorporating the same immunoreagents. Repetition of ECL assays over time and...
Crews HM, Dean JR, Ebdon L, Massey RC.The speciation of cadmium in retail pig kidney has been examined by size-exclusion chromatography (SEC) coupled directly to inductively coupled plasma mass spectrometry (ICP-MS). Approximately 35% of the cadmium from uncooked kidney was soluble after aqueous extraction at pH 8 and SEC - ICP-MS revealed three discrete peaks whose retention times corresponded to estimated relative molecular masses of 1.2 x 10(6), 7.0 x 10(4) and 6 x 10(3)-9 x 10(3). In the cooked kidney, 35% of the Cd was soluble and was all associated with a peak of a relative molecular mass (Mr) of 6 x 10(3)-9 x 10(3). After s...
Ginther OJ, Beg MA.The temporal associations of cortisol, estradiol-17β, and oxytocin with pulses of PGFM at the common hour of transition between preluteolysis and luteolysis was studied in plasma from hourly blood samples in mares (n=8). The transitional hour was determined from progesterone concentrations and occurred between 2PM and 2AM in all mares. Pulses of PGFM were grouped into those occurring at the last pulse of preluteolysis (preluteolytic pulse), at the hour of transition (transitional), and during luteolysis (luteolytic). The preluteolytic PGFM pulse (45±16pg/ml at peak) and transitional pulse (4...
Delesalle C, van Acker N, Claes P, Deprez P, de Smet I, Dewulf J, Lefebvre RA.Prokinetic drugs used to treat gastrointestinal ileus in man have equivocal results in horses. In man, prokinetic drugs have 5-hydroxytryptamine4(5-HT4) receptors as their target, but little is known about the 5-HT-receptor subtypes in the equine small intestine. Objective: Functional and immunohistochemical identification of the serotonin receptor subtype(s) responsible for the 5-HT induced contractile response in the equine circular jejunum. Methods: Isometric organ-bath recordings were carried out to assess spontaneous and drug-evoked contractile activity of equine circular jejunum. Histolo...
Guan F, Uboh CE, Soma LR, Birks EK, Teleis D, Rudy JA, Watson AO, Tsang DS.A reliable and sensitive method for the extraction and quantification of phenytoin (5,5'-diphenylhydantoin), its major metabolite, 5-(p-hydroxyphenyl)-5-phenylhydantoin (p-HPPH) and minor metabolite, 5-(m-hydroxyphenyl)-5-phenylhydantoin (m-HPPH) in horse urine and plasma is described. The method involves the use of solid-phase extraction (SPE), liquid-liquid extraction (LLE), enzyme hydrolysis (EH) and high-performance liquid chromatography (HPLC). The minor metabolite, 5-(m-hydroxyphenyl)-5-phenylhydantoin (m-HPPH) was not present in a reliably quantifiable concentration in all samples. The ...
Nakayama SM, Ikenaka Y, Hayami A, Mizukawa H, Darwish WS, Watanabe KP, Kawai YK, Ishizuka M.Research on drug metabolism and pharmacokinetics in large animal species including the horse is scarce because of the challenges in conducting in vivo studies. The metabolic reactions catalyzed by cytochrome P450s (CYPs) are central to drug pharmacokinetics. This study elucidated the characteristics of equine CYPs using diazepam (DZP) as a model compound as this drug is widely used as an anesthetic and sedative in horses, and is principally metabolized by CYPs. Diazepam metabolic activities were measured in vitro using horse and rat liver microsomes to clarify the species differences in enzy...
