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.
Fernandes I, Takehara HA, Mota I.Horse immunoglobulins were obtained from normal serum defatted with dextran sulfate and precipitated with ammonium sulfate. Eight mg of this preparation was submitted to affinity chromatography with protein A-Sepharose CL-4B. Low temperature (4 degrees C) and a starting buffer at pH 8.0 were conditions required for all IgG subclasses to bind to protein A, even those with low affinity. The IgGs bound to protein A were eluted with glycine buffer at pH 2.8. The yield was about 90%. It is suggested that isolated IgG, instead of whole Igs, be used in serum therapy, reducing the amount of Igs and di...
Penhallow RC, Mason AB, Woodworth RC.1. Human, bovine and equine transferrins have been characterized with respect to mol. wt, and behavior on urea-polyacrylamide gels, and isoelectric focussing gels. 2. As shown by SDS-polyacrylamide gel electrophoresis human transferrin has one major polypeptide whereas both bovine and equine transferrins have two polypeptides. 3. The transferrins show multiple banded patterns on urea-polyacrylamide and isoelectric focussing gels, particularly when iron saturated. The various forms are not resolved by neuraminidase treatment.
Delbeke FT, Debackere M.A high pressure liquid chromatographic (HPLC) method for measuring the theobromine content in cocoa husks, pelleted food and horse urine is described. Starting with 2 ml of urine, concentrations of 500 ng/ml could easily be detected. When feed containing 38.4 mg of theobromine was given twice daily to horses for 2 1/2 days, two days were needed after the last intake before the theobromine concentrations fell below the threshold value of 2 micrograms/ml. The time at which the peak excretion rate occurred varied from 2 to 12 h after the last administration, while the excretion rate seemed to be ...
Seither RL, Brown OR, Babu BV.Rat, cow and foal lung extracts contained an inhibitor for the liver NAD biosynthetic-pathway enzyme, nicotinatemononucleotide pyrophosphorylase (carboxylating) [EC 2.4.2.19]. The inhibitor was not dialyzable, was labile at 100 degrees C, was retained by a 30,000 dalton pore size Amicon membrane and, when partially purified by precipitation at 40-100% ammonium sulfate, inhibited the enzyme stoichiometrically. Lung reportedly does not contain nicotinate-mononucleotide pyrophosphorylase or make NAD de novo. However, the inhibitor would mask detection of the enzyme in lung extracts. We detected a...
Bhushan C, Müller I, Friedhoff KT.A rapid and simple method for concentrating leucocyte-free Babesia caballi-infected erythrocytes from in vitro cultures is described. Infected erythrocytes amounted to at least 95% of all red cells obtained.
May SA, Hooke RE, Lees P.The concentration of Percoll required for isolating equine peripheral blood mononuclear cells has been reinvestigated. A poor cell yield was obtained at the 60 per cent concentration already reported. It is recommended that workers specifically interested in high yields of mononuclear cells, for investigation of lymphocyte and monocyte functions, use a concentration of 65 per cent Percoll. However, workers wishing to isolate pure populations of equine neutrophils might consider a concentration of 70 per cent in the upper layer of Percoll used to retain the mononuclear cells.
Holtan DW, Houghton E, Silver M, Fowden AL, Ousey J, Rossdale PD.This study used gas chromatography/mass spectrometry (GC/MS) to identify and measure plasma progestagens. The method included deuterated internal standards, e.g. [17,21,21,21-2H]-5 alpha-pregnane-3,20-dione, solid-phase extraction, derivatization (methoxime/t-butyldimethylsilyl) and GC/MS. Full-scan screening identified 3 5-pregnenes, 2 4-pregnenes and 7 5 alpha-pregnanes (no 5 beta-pregnanes). The selected ion mode was used for routine quantitation from calibration curves; response was linear (r greater than 0.98) from 2 to 2000 ng equivalents/ml (0.5 ng/ml method sensitivity) and intra- and ...
Uboh CE, Rudy JA, Soma LR, Fennell M, May L, Sams R, Railing FA, Shellenberger J, Kahler M.The purpose of this study was two-fold: (1) to develop a simple and sensitive screening procedure for identifying and confirming bromhexine and ambroxol and, (2) to determine the effect of furosemide on the detection of bromhexine, ambroxol, or their metabolites in urine. Female horses (450-550 kg) treated with bromhexine or ambroxol (1 g, p.o.) were used. Urine samples were collected up to 48 h post-drug administration and analysed. Blind samples were used in evaluating the sensitivity of these methods and reproducibility of the results. Bromhexine and ambroxol were extensively metabolized in...
Bowling AT, Williams MJ.Two additional specificities (Dq and Dr) were assigned to the D system of horse red cell alloantigens following discussion of the 1989 ISAG Horse Comparison Test (HCT) results. Family and population data support 25 phenogroups defined by the enhanced battery of 17 D system factors.
Stewart F, Maher JK.The number of genes encoding the common alpha-subunit and hormone-specific beta-subunits of the equine gonadotrophins (FSH, LH and CG) were investigated in the horse (Equus caballus), donkey (E. asinus) and 2 horse x donkey hybrids (the mule and hinny). The Southern technique, involving restriction enzyme digestion, blotting and DNA hybridization to 32P-labelled DNA probes was used to estimate the copy number for each gene and to assess the extent to which equids resemble primates, the only other animals that secrete a CG during pregnancy. These methods indicated that, in common with mammals, ...
Patterson SD, Bell K, Shaw DC.1. The donkey postalbumin protein has been shown to be the equivalent of human alpha 1 B-glycoprotein by protein immunoblotting and N-terminal amino acid sequence. 2. The horse A1B system (already identified as the homologue of human alpha 1 B-glycoprotein) and the donkey alpha 1 B-glycoprotein were characterized further for terminal sialic acid content, isoelectric point, amino acid composition and affinity for the dye-ligand, Cibacron Blue F3GA (known to bind human alpha 1 B-glycoprotein). 3. Two new alleles in the horse A1B system were found, bringing the total number of alleles to five. No...
Padilla AW, Tobback C, Foote RH.A method for preparing stored unfrozen stallion spermatozoa for the zona-free hamster oocyte penetration test (HOPT) and a subsequent comparison of fresh and stored sperm by the HOPT were evaluated. In Experiment 1, sperm from 4 stallion ejaculates, cooled to 4 degrees C and stored for 24 h, were treated with 60, 90 and 120 microM of dilauroylphosphatidyl-choline (PC12) liposomes to initiate the acrosome reaction. The percentage of motile and acrosome-reacted (AR) sperm were recorded after 8, 15 and 30 min of incubation at 39 degrees C, by automated image analysis. Liposome concentration did n...
Zhang JJ, Muzs LZ, Boyle MS.Three experiments were conducted to assess the structural and functional changes of stallion spermatozoa in response to the calcium ionophore A23187, and to determine individual variation between stallions. In Experiment 1, changes in the acrosome of spermatozoa exposed to 7.14 microM A23187 for fixed times between 0 and 120 min were examined. There was a steady increase with time in the number of spermatozoa undergoing the acrosome reaction although the rate of increase differed between stallions. Sperm motility decreased sharply when incubation was extended beyond 30 min. In Experiment 2, th...
Baldwin DM, Roser JF, Muyan M, Lasley B, Dybdal N.Enzymatically dispersed anterior pituitary cells from donor mares were cultured for 48 h in alpha-modified Eagles' medium containing 10% steroid-free horse serum. The cells were then incubated for 24 h in fresh medium oestrogen followed by a 4-h incubation with or without GnRH. Media and cell extracts were analyzed for LH by radioimmunoassay. In the first series of experiments, pituitary cells from Day-3 dioestrous mares were preincubated with ethanol (control) or different concentrations of E2 (10(-11) to 10(-7) M) for 24 h prior to a 4-h incubation without (basal) or with 1.0 nM GnRH. E2 inc...
Tucker KE, Henderson KA, Duby RT.Twenty-three follicles were collected from 14 mares on specific days and grouped to represent follicles from early (Group 1; n = 6), mid (Group 2; n = 11) and late (Group 3; n = 6) oestrus, as described previously (Tucker et al., 1988). Isolated granulosa cells (GC) from each follicle were cultured in multiwell plates containing either Eagle's Minimum Essential Medium (MEM) alone, eLH (300 ng/ml), eFSH (300 ng/ml) or eLH + eFSH (300 ng/ml each), in the presence or absence of 0.5 microM testosterone. Media were collected and replaced at 24 h of culture, and 24 h later, media were again collecte...
Andrianov AM, Akhrem AA.Using the earlier suggested method the calculation of the backbone conformations of horse heart cytochrome c in oxidized (ferricytochrome c) and reduced (ferrocytochrome c) states has been performed by the two-dimensional nuclear Overhauser effect spectroscopy data. For both protein forms the secondary structure elements have been revealed and the conformations of the irregular polypeptide chain segments have been analysed. The similarity of the secondary structures of ferri- and ferrocytochrome c in solution was established from the comparison of their conformations. Small differences between...
Houghton E, Holtan D, Grainger L, Voller BE, Rossdale PD, Ousey JC.Radioimmunoassay (RIA) and gas chromatography-mass spectrometry (GC-MS) were used to determine plasma progestagen concentrations in the normal and premature foal. Radioimmunoassay provides a profile of plasma progestagens with respect to time but, due to the non-specific nature of the technique and without prior chromatographic purification, quantitative data based on RIA analysis must be interpreted with caution. In contrast, the greater specificity of GC-MS allows identification of specific plasma progestagens and measuring of multiple analytes in a single analysis. Both techniques demonstra...
Hierholzer JC, Stone YO, Broderson JR.Reference equine antisera to all 47 serotypes of human adenoviruses presently described have been prepared and evaluated by reciprocal neutralization and hemagglutination-inhibition tests. All tests were carried to endpoint dilutions a minimum of five times in each direction to give accurate values for homologous and heterologous antibody titers. Significant cross-reactions in the horse antisera were compared to similar data obtained from rabbit antisera. Using this analysis, major antigenic relationships exist among types 12-18-31 of subgenus A, types 7-11-14 and 34-35 of subgenus B, types 8-...
Angelos JA, Marti E, Lazary S, Carmichael LE.The DNA from equine sarcoid samples from New York State and Switzerland was isolated and probed with bovine papillomavirus type 1 (BPV-1) to determine if BPV genomes were present. Twelve of 13 sarcoids from New York State and 17/20 sarcoids from Switzerland contained DNA that hybridized to the BPV-1 probe. Restriction enzyme analysis of the positive samples demonstrated restriction fragment profiles characteristic of BPV-1 in 22 sarcoids and restriction fragment profiles characteristic of bovine papillomavirus type 2 (BPV-2) in 7 sarcoids. In addition, three tissues histologically diagnosed as...
Nielsen AH, Gotfredsen P, Nielsen PB, Hyttel P, Poulsen K.1. Species specific problems complicating the measurement of prorenin and renin concentrations were studied in bovine, hog and horse plasma. 2. In contrast to horse renin, bovine and hog renin reacted with rat angiotensinogen, allowing measurement of the plasma renin concentration in cattle and hog with rat angiotensinogen as exogenous substrate. 3. Trypsin treatment of plasma in order to activate prorenin generated an interfering angiotensin I immunoreactive material in all three species, most extensively in horse plasma. 4. This material could be removed in bovine and hog plasma by a cation-...
Pellegrini A, Waiblinger S, Von Fellenberg R.Lysozyme from equine neutrophil granulocytes was isolated in a pure form by fast performance liquid chromatography, i.e. ion-exchange chromatography and reversed-phase chromatography. The lysozyme lysed Micrococcus luteus, Bacillus subtilis and Staphylococcus lentus and was also bactericidal against the Gram-negative bacteria Escherichia coli, Klebsiella pneumoniae, Bordetella bronchiseptica, and Serratia marcescens. Staphylococcus aureus and Staphylococcus epidermidis were not lysed. The lysozyme was only very slightly bactericidal for S. epidermidis and S. aureus. Equine neutrophil lysozyme ...
Baker CB, Adams MH, McDowell KJ.Horse conceptuses were collected on Days 13, 15, 20 and 25 after ovulation. Whole conceptuses (Days 13 and 15) or extra-embryonic membranes (Days 20 and 25) were homogenized and poly-adenylated RNA (poly A RNA) was isolated by binding to oligo (dT)-cellulose. Poly A RNA (1 microgram/well) was separated by size on a denaturing 1% agarose gel and blotted onto nitrocellulose filters (northern blotting). DNA probes were prepared from plasmids containing equine alpha 1, omega 1 and omega 2 interferons and human beta actin. The presence of messenger RNA (mRNA) was detected by specific hybridization ...
Boermans HJ, Johnstone I, Black WD, Murphy M.Six horses gavaged with a commercial brodifacoum (BDF)-containing bait (Talone) at a dosage of 0.125 mg of BDF/kg of body weight showed weight loss, severe hypocoagulability and hemogram alterations. Four of the horses became depressed and anorectic; one required vitamin K1 therapy. Increases in clotting times were observed at 24 h in the partial thromboplastin time (PTT) followed by the thrombotest (TBT) and one-stage prothrombin time (PT) at 48 h. Elevated mean PTT, PT and TBT were observed from days 4 to 8 (p less than 0.05) with levels returning to pretreatment levels by day 12. Maximum pr...
Hermanson JW, Hegemann-Monachelli MT, Daaod MJ, LaFramboise WA.The biceps brachii of horses is subdivided into a lateral and medial head. Electrophoresis of samples from the lateral head revealed three slow-migrating native myosin isoforms, including one that does not correspond to slow myosin isoforms described for other mammalian muscles. In contrast, the medial head contained a single slow isoform. Both the lateral and medial heads contained three fast-migrating isoforms corresponding with the FM-2, FM-3 and FM-4 isoforms reported for other mammalian fast-twitch muscle fibers. Electrophoresis of myosin heavy chains (MHCs) revealed only two MHC bands, o...
Battut I, Bézard J, Palmer E.A culture for equine oviduct epithelial cells is described. Primary cultures reached confluence in 5-8 days, forming a monolayer of polygonal cells and remaining morphologically intact for about 20 days. Subcultures were obtained by collecting cells detached spontaneously from the monolayers, and confluence was reached again after 5-7 days. Cells frozen before primary culture were confluent 10-15 days after thawing. Dishes containing confluent cells also were frozen, and some cohesive monolayers formed after thawing. Equine embryos, collected 2 days after ovulation, were cultured alone or with...
McFarlane JR, Cabrera CM, Coulson SA, Papkoff H.The rhinoceros is an endangered species related to the horse family. Little is known of its reproductive endocrinology. The objectives of this study were to partially purify rhinoceros pituitary hormones, determine which assays could be used for their assessment, and to ascertain whether rhinoceros LH possesses the intrinsic FSH activity of equine LH. A single pituitary each from a White (1.3 g) and a Black (1.2 g) Rhinoceros was homogenized and extracted (pH 9.5), then subjected to pH and salt fractionation, and ion-exchange chromatography (DEAE and Sephadex SP-C50) to yield partially purifie...
Tompkins D, Hudgens E, Horohov D, Baldwin CL.This report describes the initial cloning and characterization of the equine interleukin-17 (IL-17) expressed gene sequence from mRNA obtained from equine intestinal tissue and interleukin-23 (IL-23) expressed gene sequence from mRNA obtained from equine peripheral blood mononuclear cells. Equine IL-17 has 462 nucleotides in the translated region, determined by homology with known human and mouse sequences, and shares 84% and 75% identity, respectively. For the deduced amino acid sequences, the identity with human and mouse is 76% and 70%. Equine IL-23 has 579 nucleotides in the translated reg...
Sonea IM, Wilson DV, Bowker RM, Robinson NE.Tachykinins, of which substance P (SP) is the prototype, are neuropeptides which are widely distributed in the nervous systems. In the equine gut, SP is present in enteric nerves and is a powerful constrictor of enteric muscle; in other species, SP is also known to have potent vasodilatory and pro-inflammatory effects. The specific effects of SP are determined by the subtype of receptor present in the target tissue. There are 3 known subtypes of tachykinin receptors, distinguished by their relative affinities for SP and other tachykinins. The distribution of SP binding sites in the equine pelv...
Keen B, Cawley A, Reedy B, Noble G, Loy J, Fu S.The use of catechol-O-methyltransferase inhibitors may mask doping agents, primarily levodopa, administered to racehorses and prolong the stimulating effects of dopaminergic compounds such as dopamine. It is known that 3-methoxytyramine is a metabolite of dopamine and 3-methoxytyrosine is a metabolite of levodopa thus these compounds are proposed to be potential biomarkers of interest. Previous research established a urinary threshold of 4,000 ng/mL for 3-methoxytyramine to monitor misuse of dopaminergic agents. However, there is no equivalent biomarker in plasma. To address this deficiency a...
Hurlimann J, Darling H.The synthesis of immunoglobulins by the salivary glands from eight different species was studied. It has been demonstrated that salivary glands from the cow, horse, sheep, pig, rat and guinea-pig preferentially synthesize a fast migrating immunoglobulin which seems to be analogous to IgA. In three of the species, the cow, sheep and pig, the IgA-like component cross-reacts with human IgA. The IgA synthesized by the salivary glands from the rat cross-reacts with the mouse IgA. When one compares the salivary IgA from the cow, horse, sheep, pig and rat with the IgA synthesized by the lymph nodes,...
Makinde AA.The reverse single radial immunodiffusion technique was used to detect Dermatophilus congolensis antibody in sera collected from animals previously infected to varying levels with D congolensis. Ammonium sulphate and trichloroacetic acid extracts of five different strains of D congolensis obtained from different geographical locations were used as antigens. All the extracts showed variations in their sensitivities in detecting D congolensis antibody in the various serum samples. Multiple antibodies were detected by some extracts while some showed negative antibody reaction to all extracts. Two...
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 ...
Rashmir-Raven AM, Coyne CP, Fenwick BW, Gaughan EM, Andrews GA, DeBowes RM.The ability of polysulfated glycosaminoglycans (PSGAG) to inhibit the complement cascade was evaluated. The role of complement in inflammation and infection has been well documented. Inhibition of the complement cascade by PSGAG could explain why intra-articularly administered PSGAG diminish diarthrodial joint inflammation and potentiate septic arthritis in horses. Hemolytic complement testing was performed to evaluate the effect of PSGAG on the equine classical and alternate pathways of complement, using rabbit erythrocytes as the target cells. Concentration of PSGAG between 0.2 mg/ml and 0.6...
Little CB, Hilbert BJ, Wickstrom S, Hedlund BE.An alcian blue precipitation method for quantifying the hyaluronic acid (HA) and sulphated glycosaminoglycan concentration (SGAG) in solutions containing both compounds was assessed. The assay was found to be rapid and reliable in solutions containing 0 to 200 mg of HA/dl and 50 to 1,000 micrograms of SGAG/dl, and was not affected by the presence of protein, hemoglobin, or methemoglobin in concentrations normally found in synovial fluid. The HA and SGAG concentrations in intercarpal synovial fluid from 13 clinically normal and 11 arthritic horses were evaluated. A relationship was not found be...
Qi T, Hu Y, Hu Z, Zhao S, Cullinane A, Lyons P, Gildea S, Wang X.Quantitation of virions is one of the important indexes in virological studies. To establish a sensitive and rapid quantitative detection method for equine arteritis virus (EAV), an antigen-capture enzyme-linked immunosorbent assay (AC-ELISA) was developed by using two EAV nucleoprotein monoclonal antibodies (mAbs), 2B9 and 2B3, prepared in this study. After condition optimization, mAb 2B9 was used as the capture antibody, and HRP-labeled 2B3 was chosen as the detecting antibody. The AC-ELISA had a good standard curve when viral particles of the Bucyrus EAV strain were used as a reference stan...
Corbin LJ, Blott SC, Swinburne JE, Vaudin M, Bishop SC, Woolliams JA.We have used linkage disequilibrium (LD) to identify single nucleotide polymorphisms (SNPs) on the Illumina Equine SNP50 BeadChip, which may be incorrectly positioned on the genome map. A total of 1201 Thoroughbred horses were genotyped using the Illumina Equine SNP50 BeadChip. LD was evaluated in a pairwise fashion between all autosomal SNPs, both within and across chromosomes. Filters were then applied to the data, firstly to identify SNPs that may have been mapped to the wrong chromosome and secondly to identify SNPs that may have been incorrectly positioned within chromosomes. We identifie...
Moore BD, Balasuriya UB, Nurton JP, McCollum WH, Timoney PJ, Guthrie AJ, MacLachlan NJ.To compare growth characteristics of strains of equine arteritis virus (EAV) of differing virulence to horses in rabbit kidney (RK)-13 cells and equine endothelial cells (EECs) cultured from the pulmonary artery of a foal. Methods: 13 strains of EAV, including 11 field isolates of differing virulence to horses; the highly virulent, horse-adapted Bucyrus strain; and the modified-live virus (MLV) vaccine derived from it. Methods: The growth characteristics of the 13 strains were compared in EECs and RK-13 cells. Viral nucleoprotein expression, cytopathogenicity, and plaque size were compared to ...
Higgins AJ, Lees P, Sharma SC, Taylor JB.An accurate and reliable method for the separation of flunixin from, and measurement in, equine inflammatory exudate and plasma by high performance liquid chromatography has been developed. Flunixin can be detected in concentrations as low as 0.05 micrograms/ml using an ultraviolet spectrophotometric detector at 285 nm. Samples were acidified with 2M hydrochloric acid and extracted with dichloromethane. The extract was evaporated and reconstituted in acetonitrile. Iminodibenzyl was used as internal standard. The mean recovery of flunixin from plasma was 97.6 +/- 3.9 per cent. Particular advant...
Brandon CI, Srivastava PN, Heusner GL, Fayrer-Hosken RA.Acrosin, Arysulfatase A, and beta-N-acetylglucosaminidase are three key enzymes localized within the mammalian acrosome that play a pivotal role in the penetration of the oocyte. The objectives of this study were to compare two methods of enzyme extraction based on the activities of these enzymes from equine spermatozoa. Method A utilized a 0.5 M Tris-maleate buffer containing 0.1% Triton X-100 and Hyamine 2389. Method B used 0.05 M Tris-HCl, 0.05 M MgCl2 in 0.05 M Tris-maleate, followed by 0.05 M Tris-maleate containing 0.1% Triton X-100. Results indicated that acrosin was initially bound in ...
de Bruijn CM, Wensing T, van Nieuwstadt RA.The glutaraldehyde coagulation test is a semi-quantitative test used to determine the gammaglobulin concentration in serum. The purpose of this study was to examine the reliability of the different modifications of this test by determining the sensitivity, specificity, positive and negative predictive value, and the prevalence of hypogammaglobulinemia in foals. The results of the glutaraldehyde coagulation test were compared with the serum gammaglobulin concentration as a reference value, determined by measuring total serum protein and the serum protein spectrum. It was concluded that the glut...
Aguilar JJ, Woods GL, Miragaya MH, Olsen LM.The object of this experiment was to estimate the number and type of living cells in oviductal masses of mares. Oviducts of abattoir mares were dissected, divided into 3 sections, and flushed individually. Oviductal masses were recovered from 220 of 250 mares and from 389 of 500 oviducts. A greater number of masses was recovered from the left than the right oviducts. A higher percentage of masses was recovered from the ampullary-isthmic junction than from the ampulla or isthmus. The number of masses increased slightly with increasing mare age and was weakly correlated with the number of unfert...
Messier PE, Drouin R, Richer CL.We present an immunochemical technique for the detection of 5-bromo-2'-deoxyuridine (BrdU) incorporated discontinuously into the chromosomal DNA. A monoclonal anti-BrdU antibody and a protein A-gold complex were used to produce chromosome banding of human and equine chromosomes, specific for electron microscopy (EM). Well-defined bands, symmetry of sister chromatids, concordance between homologues, and band patterns similar to those observed by light microscopy facilitate chromosome identification and karyotyping. From prophase to late metaphase, chromosomes condense and bands appear to fuse. ...
Caloni F, Spotti M, Villa R, Mariani C, Montana M, Pompa G.An investigation was undertaken to demonstrate whether therapeutic treatment with ACTH raises hydrocortisone (cortisol) levels in horse urine above the limit (1000 ng/ml) established by the International Conference of Racing Authorities with the aim of controlling the abuse of cortisol and ACTH in equine sports. ACTH (200 iu) was administered i.m. to 3 Thoroughbred horses; urine and blood samples were collected at intervals afterwards and analysed by an immunoenzymatic system (ELISA) and HPLC-MS. To ascertain post exercise cortisol levels in untreated horses, 101 urine and 103 serum samples we...
Luque I, Fernández-Garayzábal JF, Blume V, Maldonado A, Astorga R, Tarradas C.The anti-microbial susceptibility and genetic diversity of 65 strains of Streptococcus equi ssp. zooepidemicus (Sez) isolated from mares presenting clinical signs of endometritis was determined by disk agar diffusion and pulsed field gel electrophoresis (PFGE) methods, respectively. Overall, Sez isolates were susceptible to beta-lactams, enrofloxacin, trimethoprim-sulfamethoxazole and gentamicin. These anti-microbials could be recommended as empiric anti-microbial therapy in cases of endometritis caused by Sez. Pulsed field gel electrophoresis typing revealed a great genetic diversity (56 diff...
Bastos FZ, Barussi FCM, Santi TF, Vieira BP, Senegaglia AC, Cruz FF, Michelotto PV.There is no consensus on aspects of equine bone marrow collection and processing. The study aimed to describe the collection of large volumes of bone marrow from horses of advanced age, with emphasis on bone marrow mononuclear cells (BMMCs) recovery and viability after cryopreservation. Fourteen horses, aged 3-24 years, were divided into three experiments. E1 studied the feasibility of collecting 200 mL from the sternums of horses of advanced age; E2 examined the number of cells obtained from the first and last syringe of each puncture; and E3 investigated the influence of heparin concentrati...
Hall JA, Hoyt D, Zuver C, Skinner MM, Schlipf JW.The criteria used to diagnose recurrent airway obstruction (RAO) in affected horses include demonstration of reversible lower airway obstruction and greater than 25% neutrophils in bronchoalveolar lavage fluid (BALF). Additional objective laboratory tests are needed to improve diagnostic accuracy and to monitor response to treatment. The goal of this study was to determine if neutrophil chemoattractant activity of BALF could be measured by using a previously described, rapid, multiwell colorimetric assay for chemotaxis. In this assay, neutrophils that have migrated through a membrane filter ar...
Kumar J, Chaudhury A, Yadav SC.Trypanosomosis (Surra) is an economically important disease caused by Trypanosoma evansi which is an extracellular parasite present in the plasma, tissues and other body fluids of a wide range of hosts including domesticated animals. Currently, serological reports are based on detection of antibodies by ELISA using whole cell lysate (WCL) antigen, which has a limitation of persistence of anti-trypanosomal antibodies after successful treatment of the disease. Moreover, it has some ethical issues also like requirement of mice for in vivo maintenance of parasite for preparing the antigen. Therefo...
Creel JA, Stover SM, Martin RB, Fyhrie DP, Hazelwood SJ, Gibeling JC.The compliance technique has been used to monitor crack length during fracture and fatigue testing of materials. Difficulties arise when this technique is applied to anisotropic biological materials such as bone. In this tutorial, two different methods of analyzing compliance calibration data are described: the standard ASTM method and a new approach developed by the authors specifically for anisotropic materials. An example is given showing how data from equine cortical bone can be analyzed. In this example, calibration tests were conducted on thirty-six three point bend specimens machined fr...
Gil L, Saura S, Echegaray A, Martinez F, de Blas I, Akourki A, Gonzalez N, Espinosa E, Josa A.The present study evaluated the effect of supplementing the medium used to mature equine oocytes in vitro with oestrous mare serum (EMS) or horse follicular fluid (HFF). To this end, 144 ovaries were obtained from mares aged 16-21 months and transported to the laboratory in Dulbecco's phosphate buffered saline (D-PBS) at 30 degrees C. Oocytes were harvested from the ovaries by slicing, and then selected for in vitro maturation (IVM) according to the number of cumulus cell layers and the characteristics of the cytoplasm. The selected oocytes were washed three times in TCM199 medium plus HEPES (...
Woods WE, Wang CJ, Houtz PK, Tai HH, Wood T, Weckman TJ, Yang JM, Chang SL, Blake JW, Tobin T.A one step enzyme-linked immunosorbent assay (ELISA) and a particle concentration fluorescent immunoassay (PCFIA) test for furosemide were evaluated as part of a panel of pre- and post-race tests for illegal medication of racing horses. These tests are very sensitive to furosemide with an I-50 for furosemide of about 20 ng/ml. The test is also rapid; an average pre-race complement of 10 samples can be analyzed in 90 minutes or less. The ELISA test results can be read with an inexpensive spectrophotometer, or even by eye. Both the PCFIA test and the ELISA test readily detect the presence of fur...
Zhang CH, Grünig G, Davis W, Antczak DF.The regulatory effects of phorbol myristate acetate (PMA) on the expression of the CD4 molecule on horse T cells were investigated. On both peripheral blood lymphocytes and thymocytes, PMA resulted in a rapid and transient down-regulation of equine CD4 expression, but had no such effect on the surface expression of equine CD5, CD8 or major histocompatibility complex (MHC) class I and class II molecules. Over 75% of the surface CD4 molecules per cell were lost after a 4 h exposure to PMA at 37 degrees C. The regulation of equine CD4 expression induced by PMA was temperature dependent and revers...
Ferrante PL, Kronfeld DS.Collection of a satisfactory blood sample requires special procedures to prevent changes in glucose and lactate content after the sample has been obtained. Changes in measured plasma glucose and blood lactate concentrations attributable to anticoagulants and storage procedures, respectively, were examined in blood samples obtained from horses at rest and after exercise. To evaluate the effect of anticoagulants on measured plasma glucose concentration, blood was preserved with either sodium fluoride/potassium oxalate or lithium heparin. Measured plasma glucose concentration in blood obtained at...
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...
Rahnama S, Vathsangam N, Spence R, Anderson ST, de Laat MA, Bailey S, Sillence MN.Prolonged hyperinsulinemia is thought to be the cause of equine endocrinopathic laminitis, a common and crippling disease of the foot, for which there are no pharmacologic treatments other than pain relief. It has been suggested that insulin causes its effects on the lamellae by activating IGF-1 receptors (IGF-1R), as insulin receptors (InsR) are scarce in this tissue, whereas IGF-1R are abundant and become downregulated after prolonged insulin infusion. As a first step toward confirming this mechanism and beginning to develop a therapeutic anti-IGF-1R monoclonal antibody (mAb) for horses, it ...
Carstanjen B, Hoyle NR, Gabriel A, Hars O, Sandersen C, Amory H, Remy B.To evaluate a human assay for quantification of carboxy-terminal cross-linking telopeptide of type I collagen (CTX-I), assess the influence of age on plasma CTX-I concentration, investigate the relationship between plasma CTX-I and serum osteocalcin concentrations, and determine whether concentrations of plasma CTX-I or serum osteocalcin fluctuate in circadian manner in horses. HORSES: 75 clinically normal horses. Methods: Cross-reactivity between equine serum CTX-I and CTX-I antibodies in an automated electrochemiluminescent sandwich antibody assay (ECLIA) was evaluated via a specificity test...
Brook D.A guarded, sterile swab is used to obtain samples for uterine culture. With the mare in stocks, the tail bandage and the perineum washed, the culture rod is introduced into the vagina with a gloved hand. After the rod is guided through the cervix, the guard cap is dislodged and the swab is rubbed along the endometrium, after which the rod is extracted. Samples for uterine culture should only be obtained during full estrus. Swabs should be directly plated onto agar within 2 hours of collection. Blood agar is appropriate for initial screening, but use of specialized types of agar expedites ident...
