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.
Holland RA.Using a double beam stopped-flow apparatus, measurements were made of the velocity constant of the reaction CO + Hb --> COHb in solution and in the red cells of human beings, rabbits, horses, and goats. The solution constant (l') at 37 degrees C for human beings was 362 mM(-1) sec.(-1); in other species l' was somewhat lower. Two rabbits, despite having apparently identical hemoglobins had significantly different values for l'. The energy of activation (E) of l' was between 8 and 11 kcal/mole in all cases. The cell reaction constant (l'(c)) at 37 degrees was between 61 and 73 mM(-1) sec.(-1...
Black JA, Leaf G.1. The effects of cyanogen bromide on horse-heart cytochrome c and horse-heart myoglobin have been investigated. Cytochrome c yielded four fragments, of which two were haemopeptides. The two colourless peptides had amino acid compositions corresponding to those that are expected, on the basis of the sequence proposed for horse-heart cytochrome c by Margoliash, Smith, Kreil & Tuppy (1961), from cleavage at both methionine residues. Of the two haemopeptides, one was isolated and shown to be that derived from cleavage at only one methionine residue, that nearer to the C-terminus of the peptid...
Thompson NE, Bergdoll MS, Pattee PA.The factor in normal sera primarily responsible for the enhancement of transfection (and transformation) of Staphylococcus aureus was identified as fibronectin. Serum samples which were depleted of fibronectin by affinity chromatography showed a marked decrease in enhancing activity. Fibronectin isolated from sera of several animal species demonstrated enhancing activity.
Raeside JI, Christie HL, Betteridge KJ.Steroid production and metabolism by early conceptuses are very important for the establishment and maintenance of pregnancy in horses. Our earlier work suggested the possible formation of 5alpha-reduced steroids in equine conceptuses. We have now demonstrated the formation of 5alpha-reduced metabolites of androstenedione, testosterone, and progesterone by the embryo and its membranes. A total of 44 conceptuses were collected from 26 mares between 20 and 31 days of pregnancy. Tissues from the embryo proper and from the separated components of the conceptus (bilaminar and trilaminar trophoblast...
Chen JW, Uboh CE, Robinson MA, Jiang Z, Soma LR.To evaluate plasma interleukin 6 (IL-6) concentration in Standardbred racehorses by means of a novel ELISA following validation of the assay for use with equine plasma samples. Methods: Plasma samples obtained from 25 Thoroughbreds for use in assay validation and from 319 Standardbred racehorses at rest 2 to 2.5 hours prior to warm-up and racing. Methods: A sandwich ELISA was developed with equine anti-IL-6 polyclonal antibody and the biotin-streptavidin chemical interaction to enhance sensitivity. The assay was validated for specificity, sensitivity, precision, and accuracy by use of both rec...
Björkman C.1. Comparative data are presented of the activities of pyruvate dehydrogenase complex and acetyl-CoA synthetase and of the acetate content in homogenates from ventral grey matter in spinal cord from cows and two non-ruminant species, pigs and horses. The methods used in the study are evaluated and discussed. 2. The total pyruvate dehydrogenase complex activity was 24.9-29.9 mU/mg protein and did not differ between the species. The part of the complex that was in active form at the sampling occasion was 60, 85 and 95% in cows, pigs and horses, respectively. 3. Acetyl-CoA synthetase activity dif...
Bosken JM, Lehner AF, Hughes CG, Woods WE, Camargo FC, Harkins JD, Boyles J, Tobin T.Isoxsuprine is used to treat navicular disease and other lower-limb problems in the horse. Isoxsuprine is regulated as a class 4 compound by the Association of Racing Commissioners, International (ARCI) and, thus, requires regulatory monitoring. A gas chromatography-mass spectrometry method utilizing electron impact ionization was developed and validated for the quantitation of isoxsuprine in equine plasma or equine urine. The method utilized robotic solid-phase extraction and tri-methyl silyl ether products of derivatization. Products were bis-trimethylsilyl (TMS) isoxsuprine and tris-TMS rit...
Ledwozyw A, Kadziołka A, Jabłonka S, Tusińska E, Herbut M.The aim of this study was to investigate the platelet factor 3 activity in platelet-rich plasma of horse, cattle, sheep and pig, by the use of chromogenic tripeptide substrate H-D-Phenylalanyl-Pipecolyl-Arginyl-p-nitroanilide. Among species examined the highest activity of this factor was stated in pig, the lowest one in sheep. Chromogenic substrate test was 10 times more sensitive that Stypven clotting time test. Thus, the use of chromogenic tripeptide substrate is fully valuable in platelet factor 3 activity estimation in domestic animals.
Haysom LZ, Lee-Fowler TM, Spangler EA.Storage and temperature significantly impact bronchoalveolar lavage fluid (BALF) analysis, and shipment of samples to diagnostic laboratories is often necessary. Alternative sample preparation methods could limit storage and temperature effects. Objective: This study aimed to determine if airway wash samples that were fixed in formalin after being embedded in Histogel or Gelfoam gave comparable results to fresh cytocentrifuged or sediment smear preparations for the evaluation of cell morphology. Methods: Eleven bronchoalveolar lavage and 3 transtracheal wash fluids were available, including 8 ...
Rotkina AS, Pronina IV, Lazarev VN, Akhaev DN, Baskova IP.The purpose. Identifying the capacity of the medicinal leech novel original recombinant thrombolytic preparation Destabilase-Lysozyme-2 to inhibit the blood platelet aggregation. Gene of destabilase-lysozyme. ds2 (mlDL-Ds2 ), was cloned in E.coli cells. Recombinant protein was isolated in denaturing conditions using metal-chelate chromatography followed by denaturation of the polypeptide by rapid dilution in exact accordance with the procedure described by Kurdyumov A.S. et al. ( 2016, Russian Journal of Bioorganic Chemistry, v.42, s. 42-52). Blood was collected from the jugular vein of 18 hor...
Bachmann V, von Salis B, Fürst A.The goal of this study was to describe the development of equine drug testing in horses in Switzerland. This was achieved through evaluation of a film made by the Institute of Forensic Medicine at the University of Basel entitled 'Doping von Rennpferden' [Doping of Race Horses], toxicological detection, 1962', the analysis of doping test results of the Swiss Equestrian Federation and by interviewing individuals of various professions who were involved in equine drug testing at the time. The study compares early and modern methods of drug testing and highlights the changes in the attitude of eq...
Wells M, Gaffin SL.Passive immunotherapy with anti-lipopolysaccharide hyperimmune equine plasma (Anti-LPS) is effective in treating experimental Gram-negative bacterial infections. The bactericidal activity of anti-LPS towards five different Pseudomonas species, including two multiresistant Pseudomonas aeruginosa isolates was tested here, as well as the ability of anti-LPS to inhibit the quantitative chromogenic limulus amoebocyte lysate (LAL) assay. Anti-LPS caused a mean reduction of 84.4 +/- 3.2% (P less than 0.001) in the number of colony forming units (cfu) of all isolates, whereas saline and complement ina...
Metenier L, Grosclaude J, Meriaux JC.The chief application of blood typing in domestic animals is in the verification of parentage. However, the acquisition of good standardized reagents in sufficient quantity remains an obstacle for the development of this work. The production of monoclonal antibodies directed against blood group determinants offers an attractive means of improving both the quality and quantity of serological reagents, and could facilitate the definition of new specificities. Fusions between a mouse myeloma line and splenocytes from mice immunized with horse red cells have resulted in four hybridomas producing a...
Theodorakis JL, Armes LG, Margoliash E.beta-Thiopropionyl derivatives of horse cytochrome c singly modified at each of 18 different lysine epsilon-amino groups have been prepared using sulfosuccinimidyl-2-(biotinamido)ethyl-1,3-dithiopropionate and purified to homogeneity by high-pressure liquid chromatography. These derivatives were characterized by determination of: (i) the location of the modification; (ii) reduction potentials; (iii) visible and NMR spectra: and by (iv) measurement of electron transfer activity with cytochrome-c oxidase. No significant changes in structure were indicated, except for the ferric forms of the deri...
Kasza A, Korpula-Mastalerz R, Rose-John S, Dubin A.The horse leucocyte elastase inhibitor (HLEI), present in neutrophils, monocytes and bone marrow cells, is apparently a cytoplasmic protein which is not released from cells even in response to stimulation with lipopolysaccharide, phorbol ester, tumour necrosis factor alpha, interleukin-1 or elastin degradation products. Although no expression of the inhibitor was detected in neutrophils, both monocytes and bone marrow cells were efficient in its synthesis. Using a new expression vector pREST5d, recombinant inhibitor was produced in a large quantity in a soluble form, with a yield of 88 mg per ...
Franken P, Schotman JH.The activities and concentrations of a number of erythrocytic enzymes and intermediate products of erythrocyte metabolism were determined in twenty-one normal standard-bred horses which were studied clinically and biochemically. These studies showed that equine anaerobic glycolysis is characterized by a biochemical pattern similar to that observed in human PK deficiency. The greater sensitivity of equine haemoglobin to oxidants is attributable either to low stability of GSH, which may be due either to the low activity of GR or that of 6PGD as observed in the studies. In addition, the saturatio...
Zielińska-Dawidziak M, Błaszak R, Piasecka-Kwiatkowska D.The presented experiments show the model of expectation of equine spleen ferritin extraction in a new aqueous two-phase system which was formed by mixing polyethylene glycol (PEG) and hydroxyethyl starch (HES). The tendency of the protein to migrate in the analyzed systems was dependent on the concentrations of HES and PEG as well as PEG molecular weight. The highest concentration of ferritin in the top phase (rich in PEG) was recorded in the system composed of 6% PEG 3000 and 3% HES. The obtained concentration was 0.88 mg mL(-1). The lowest concentration was 0.42 mg mL(-1) in the system compo...
Shivers CA, Liu IK.Sera and follicular fluid from mares previously determined as having antibodies to the zona pellucida were exposed to porcine ovarian oocytes before insemination with boar spermatozoa in vitro. There was a dramatic decline in the number of spermatozoa bound to zonae compared to treatments with sera and follicular fluid from mares negative for zona antibodies. These data suggest that antibodies reactive with the zona pellucida may be responsible for reduced or even complete infertility in some mares. In a group of 50 randomly selected mares tested for antizona antibodies, 2 pregnant mares were ...
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...
Eichhorn W.Infections and clinical diseases caused by equine 2 influenza A viruses are observed worldwide. The frequency of these outbreaks supports the hypothesis that antigenic variation of the surface proteins may play an important role. For the demonstration of these variations, monoclonal antibodies (Mabs) were prepared. They are directed against the hemagglutinin or the neuraminidase of the prototype strain a/eq/Miami/1/63. In hemagglutination-inhibition assays with Mabs two reaction patterns were observed: four Mabs inhibited 14 out of 17 strains tested. Another Mab recognized the hemagglutinin of...
Word TA, Larsen RW.Treatment of horse heart Cytochrome-c (Cc) with N-chloro-4-toluosulfonamide (Chloramine-t, CT) results in the oxidation of methionine (Met) residues to the corresponding sulfoxide including the distal heme ligand, Met80. The resulting Fe-sulfoxide coordination is sufficiently labile in the ferrous form to be displaced by gaseous ligands, including CO. Photolysis of the CO-CT-Cc complex provides an opportunity to examine ligand binding dynamics that are associated with a relatively rigid distal heme pocket. In this work, photoacoustic calorimetry (PAC) was utilized to obtain the kinetics as wel...
Leblová S, Galociová J, Cerovská N.The studied herbicides (terbutylazine, simazine) inhibit the activity of plant, animal, and yeast alcohol dehydrogenases. The inhibition constant Ki for alcohol dehydrogenase (ADH) isolated from peas and bakers' yeast equals approximately 10(-4) M, and that for ADH isolated from horse liver is of the order of 10(-5) M. The character of inhibition for all the herbicides studied for the reaction catalyzed by pea, liver, and yeast ADH is always noncompetitive toward ethanol and competitive with respect to NAD. The inhibition constants for the enzyme isolated from peas are pH independent. The inte...
Reglero A, Hueso P, Rodrigo M, García-Alonso J, Llanillo M, Cabezas JA.The ganglioside patterns of the forebrain, cerebellum and brain stem from horse, donkey, mule and goat have been determined by thin-layer chromatography. GM1, GD1a, GD1b and GT1b are the four major brain gangliosides. N-acetylneuraminic acid as the predominant sialic acid (congruent to 97%) and traces of N-glycolyneuraminic acid were found. The four above mentioned major gangliosides were also found in the forebrain, cerebellum and brain stem of adult rats. This pattern is not modified in rats under stress situation (at 4 degrees C for 3 months). In other extraneural organs from rats such as l...
Gupta AK, Kaur D, Rattan B, Yadav MP.Three abortigenic Indian isolates of equine herpesvirus-1 (EHV-1) (Tohana, Hisar and Bikaner), along with two exotic abortigenic isolates (AB4 and V592) and another EHV-1 isolate (Jind) obtained from a case of perinatal foal mortality, were studied for variability. For this purpose, PCR and restriction endonuclease (RE) digestion techniques were used simultaneously as a DNA fingerprinting system. Nine different regions of EHV-1 virus were amplified by PCR using primer pairs specific for the regions and the products obtained from these regions were subsequently subjected to various restriction ...
Sahmi F, Nicola E, Price CA.The objective was to establish a cell line-based bioassay for FSH in horse serum for screening samples with high eCG bioactivity. A cell line (HEK293) was transiently cotransfected with an FSH reporter expression plasmid and a cAMP-responsive β-galactosidase reporter plasmid. Cells were bulk frozen, and thawed for assay purposes. This assay was specific for FSH, with no cross-reaction with LH or insulin-like growth factor-1. Standard curves (eCG) and serum samples from pregnant mares passed parallel line bioassay validity tests (linearity and parallelism). Estimates of bioactivity with this b...
Nazmutdinov RR, Bronshtein MD, Zinkicheva TT, Hansen NS, Zhang J, Ulstrup J.Outer-sphere electron transfer (ET) between optically active transition-metal complexes and either other transition-metal complexes or metalloproteins is a prototype reaction for kinetic chirality. Chirality as the ratio between bimolecular rate constants of two enantiomers mostly amounts to 1.05-1.2 with either the Λ or Δ form the more reactive, but the origin of chirality in ET parameters such as work terms, electronic transmission coefficient, and nuclear reorganization free energy has not been addressed. We report a study of ET between the Λ-/Δ-[Co(Ox)3](3-) pair (Ox = oxalate) and hor...
Breuil MF, Joseph M, Petry S.The gold standard method to isolate and identify Taylorella equigenitalis, the contagious agent of equine metritis, is the culture method according to the World Organisation for Animal Health Terrestrial Manual. No selective T. equigenitalis chocolate agar medium has been developed since the 1980s and the existing media show limited performances due to the fastidious nature of T. equigenitalis and the presence of interfering bacteria in the genital tract of equines. Here, the growth rates of 6 T. equigenitalis strains and 7 non-T. equigenitalis strains were compared on Timoney's selective medi...
Sugo T, Kato H, Iwanaga S, Fujii S.On incubation of purified horse plasma high-molecular-weight kininogen with purified plasma kallikrein, three new peptides, named fragment 1.2, fragment 1 and fragment 2, were released, in addition to the vasopeptide, bradykinin. Fragment 2 contained an extremely high level of histidine, in which eleven residues out of the total 48 residues were characterized. Thus the result proves the existence of the histidine-rich region in horse high-molecular-weight kininogen, which is similar to the region previously identified in bovine high-molecular-weight kininogen. Moreover, we have identified a ne...
Wang Y, Du X.The miscibility and stability of the binary monolayers of zwitterionic dipalmitoylphosphatidylcholine (DPPC) and cationic dioctadecyldimethylammonium bromide (DOMA) at the air-water interface and the interaction of ferritin with the immobilized monolayers have been studied in detail using surface pressure-area isotherms and surface plasmon resonance technique, respectively. The surface pressure-area isotherms indicated that the binary monolayers of DPPC and DOMA at the air-water interface were miscible and more stable than the monolayers of the two individual components. The surface plasmon re...
Qi T, Wang X.Reverse genetics is one of the most powerful tools in modern virology. Equine arteritis virus (EAV) is the prototype member of the Equartevirus. In this study, a new reverse genetics system for the recovery of equine arteritis virus from a cDNA plasmid, which contains viral cDNA sequence flanked by hammerhead ribozyme (HamRz) and hepatitis delta virus ribozyme (HdvRz) sequences in both terminals of the viral genome, was developed by optimization of the promoter and terminator regions. Cellular RNA polymerase II drove the transcription of the viral genome. The results showed that the rescued vi...
Pérez-Castrillo S, González-Fernández ML, Gutiérrez-Velasco L, Villar-Suárez V.To characterize the ultrastructure of mesenchymal stem cells (MSCs) that were harvested from the adipose tissue (AT-MSCs) and bone marrow (BM-MSCs) of horses and transfected with green fluorescent protein. Methods: MSCs from adipose tissue and bone marrow of 6 adult female Hispano-Bretón horses. Methods: Harvested equine MSCs were cultivated and transfected with green fluorescent protein, and the immunophenotypes of the MSCs were characterized by use of anti-CD90 and anti-CD105 monoclonal antibodies. When stable transfection of MSCs was achieved, the morphological and ultrastructural characte...
Dyson DA, Smith GR.When incorporated in solid medium at a concentration of 15 per cent, the defibrinated blood of certain horses strongly suppressed the growth of some, but not all, strains of Mycoplasma mycoides subsp mycoides so that many colonies failed to develop to a visible size. Blood from a single rabbit was tested and found to exert a similar effect. There was striking variation in the degree of inhibition produced by different samples of horse blood and, of five strains of the organism examined, the T1 vaccine strain was the most susceptible. The results suggested that the effect was not due to antibod...
