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.
Steinhardt J, Hiremath CB.Many of the stability characteristics of horse ferrihemo-globin (Hb+) in acid solutions, such as pH dependence and susceptibility to stabilization by iron ligands, are shared by human ferrihemoglobin, but striking differences between the two proteins exist. The most noticeable is the much greater rate of denaturation of the human protein at all pH values. Other differences include a shift to higher pH in the equi-librium between native and acid-denatured forms, differ-ences in the temperature at which the temperature effect on the equilibrium-pH curve reverses, a complete absence in human Hb+ ...
Rogers HJ.Under a variety of conditions of concentration, Eh, and pH, horse anti- serum and normal horse serum exerted similar bacteriostatic effects against Type A. Ferric iron abolished the bacteriostatic effect when added during the first 2 hours of incubation at Eh+60 mV. Ferrous iron abolished the bacteriostatic effect when added after 3 hours. Ferric iron abolished the bacteriostatic effect at—140 mV. A mixture consisting of horse β- and γ-globulins together with human transferrin exerted a bacteriostatic effect similar to that of whole serum. This system responded in the same way as whole se...
Rockey JH.Eight antigenically unique immunoglobulins have been identified in purified equine anti-p-azophenyl-beta-lactoside (Lac) antibody isolated from a single horse. The Fc fragments of the gammaGa-, gammaGb-, gammaGc-, and -gammaA-globulins have been shown to possess unique antigenic determinants. Common gammaG- and gammaA-Fc fragment antigenic determinants, which were absent from the 10Sgamma(1)- and gammaM-globulins, have also been observed. All antibody populations share two antigenically distinct light (B, L) chain variants. The association of anti-Lac antibody with the hapten p-(p-dimethylamin...
Haque RU.A technique for identifying and characterizing staphylococcal hemolysins by first separating them electrophoretically in barbital-buffered agar gel (pH 8.4) at 5 ma/cm for 2 hr and then determining their hemolytic activities by exposing them to human, horse, rabbit, and sheep erythrocytes is described. The alpha-hemolysin produced by a White variant of the Wood 46 strain of Staphylococcus aureus migrated 18 mm towards the cathode, and it lysed horse, rabbit, and sheep erythrocytes, whereas a Clear variant of the Wood 46 strain of S. aureus produced a lysin which migrated similarly to the alpha...
Irfan M.Studies have been made on the effect of storage and temperature on factor V in animals and a method for its assay is described. There was a decrease of the factor with age in oxalated plasma of cattle, horses, sheep, dogs, rabbits and one elephant. This deterioration was very rapid in plasma kept at 37C. as compared with that stored at 4C. Rabbit plasma had the highest concentration of factor V followed by dogs, horses, cattle and sheep. Mixing various dilutions of aged plasma kept at 4C. wlth fresh plasma would enable the percentage prothrombin activity to be determined for the assay of facto...
Böttiger LE.Erythrocyte sedimentation rate, total protein and fibrinogen, electrophoretic protein pattern, and total serum protein-bound carbohydrates have been determined in a number of domestic animals and compared to human values. The striking finding is that although the E.S.R. varies widely between various species, the fibrinogen content is of the same order of magnitude in all. The horse, which shows a very high E.S.R., has a well marked beta-globulin fraction as an outstanding feature, a finding that should be further studied. Blutsenkungsgeschwindigkeit, Gesamteiweiss und Fibrinogen, elektroforeti...
Broström H, Hellström U, Hammarström S, Obel N, Perlmann P.Untreated and neuraminidase-treated equine peripheral blood lymphocytes were analysed for binding of the A hemagglutinin of the snail Helix pomatia (HP). For optimal staining by direct immunofluorescence, the concentration of neuraminidase had to be increased as compared to that needed for other species. Moreover, higher concentrations of HP were required for optimal staining of equine lymphocytes as compared to lymphocytes from other species. Even so, the maximal number of equine lymphocytes exhibiting positive staining was only about 20%. No, or very few, HP-positive lymphocytes were seen wh...
Cerva L, Cerná Z.A description is given of the preparation of antigen from Sarcocystis dispersa cystozoites and the procedure of the indirect haemagglutination test (IHA). The antibodies against this antigen were detected in experimentally infected mice from day 20 p.i. (1: 640). In the following weeks the antibody titres reached the value of 1: 40,960. The sera of pigs, sheep and horses spontaneously infected with other Sarcocystis species reacted with this antigen in low titres only. The bovine sera gave negative reactions even in cases when Sarcocystis cysts were present in the muscles of the examined anima...
Wang X, Kole AR, Greenwald GS.This study was designed to determine whether the major site of eCG neutralization by an antiserum to the hormone is at the peripheral or ovarian level. Hamsters hypophysectomized at oestrus were injected s.c. with 25 iu eCG. Three days later, preovulatory follicles were dissected and cultured for 5 h and the medium was changed every hour. At the end of the first hour of incubation, oestradiol and androstenedione accumulation was high, with a sharp drop over the next 4 h, whereas progesterone concentrations did not change over the entire period. Addition of eCG antiserum to the incubated follic...
Van Beek H, Baars AJ.Metallothioneins (MTs) were separated and quantitated by reversed-phase high-performance liquid chromatography (RP-HPLC), in direct combination with atomic absorption spectrometry (AAS) for quantitation of the metal contents in MTs. MTs were eluted from an RP-8 column with a gradient of Tris buffer pH 7.0 and methanol, and were detected by UV absorbance (220 nm). Commercially available purified MTs from horse kidney and rabbit liver were analyzed for purity and metal composition. One lot of horse kidney yielded only 50% of the estimated value. In some cases, the certified metal content differe...
de Val N, Herschbach H, Potier N, Dorsselaer AV, Crichton RR.An essential difference between eukaryotic ferritins and bacterioferritins is that the latter contain naturally, in vivo haem as Fe-protoporphyrin IX. This haem is located in a hydrophobic pocket along the 2-fold symmetry axes and is liganded by two Met 52. However, in in vivo studies, a cofactor has been isolated in horse spleen apoferritin similar to protoporphyrin IX; in in vitro experiments, it has been shown that horse spleen apoferritin is able to interact with haem. Studies of haemin (Fe(III)-PPIX) incorporation into horse spleen apoferritin have been carried out, which show that the me...
Rice DW, Blake CC.Crystals of horse muscle 3-phosphoglycerate kinase have been grown in the presence of a wide variety of substrates using either potassium tartrate or polyethylene glycol as a precipitant. In those grown from polyethylene glycol, two related crystal forms have been obtained by varying the nature of the substrates present in the crystallization medium. In order to obtain one of these forms, form B, the presence of the substrate 3-phosphoglycerate appears to be essential. The two crystal forms are not interconvertible by simple diffusion experiments and the crystals grown in the absence of 3-phos...
Broström H, Obel N, Perlmann P.Human lymphocytes displayed a frequent natural cytotoxicity (NK) in vitro against normal equine dermal fibroblasts (ED) and against equine tumour cells of a virus-containing cell line (Mc-1). Similarly, human normal sera contained antibodies that induced antibody-dependent cellular cytotoxicity (ADCC) by normal human lymphocytes against the same target cells. Both NK and ADCC varied for different donors. For individual donors, however, cytotoxicity against the two target cells was significantly correlated both in NK and ADCC. For ED there was also a significant correlation between ADCC and NK ...
Neuwirth L, Romine C.The construction of ancillary equipment used to improve image quality and reduce personnel radiation exposure in the equine nuclear medicine laboratory is illustrated. The devices include a self supporting lead sheet for shielding the distal limb or limb pair, a hanging lead sheet for shielding the proximal limb, a lead square for shielding the urinary bladder or jugular catheter, a restraining board for acquiring a palmar view of the foot, a head support to stabilize the head for imaging and a head support for stabilizing the neck for imaging. The restraining board and head supports decrease ...
Alben JO, Bare GH.Infrared absorption spectra of the alpha-104 (G11) cysteine SH group have been observed for aqueous solutions of hemoglobin derivatives from humans, pigs, and horses. The center frequencies ((nu)SH) show ligand sensitive patterns that are similar for the three species, with (nu)SH (HbCO) <(nu)SH (HbO(2) ~ HbCN) < (nu)SH (Hb(+)) <<(nu)SH (deoxyHb) for human and pig hemoglobins. The alpha-104 SH group is most strongly H-bonded (smallest (nu)SH), has the greatest range of (nu)SH (Hb ? HbCO) in human hemoglobin, and is least strongly H-bonded and has the smallest range of (nu)SH (Hb ? HbCO) in hor...
Woods WE, Chay S, Houston T, Blake JW, Tobin T.Interference or 'masking' in thin layer chromatography occurs when the presence of one drug on a thin layer plate physically obscures or interferes with the detection of another drug. We investigated the ability of phenylbutazone and oxyphenbutazone to mask or interfere with the detection by high performance thin layer chromatography (HPTLC) of basic drugs used illegally in horse racing. Of fifty-five basic drugs called 'positive' since 1981 by laboratories affiliated with the Association of Official Racing Chemists (AORC), forty did not comigrate with phenylbutazone or oxyphenbutazone and cou...
Xu J, Cai J, Suresh M, Peek SF, Darien BJ.Equine PSGL-1 (ePSGL-1) is widely expressed on equine PBMC as a homodimer with sialylation (sLeX) modifications that contribute to P-selectin binding affinity. To investigate the role of other potential post-translational modifications required for high-affinity P-selectin binding, ePSGL-1 was transfected into CHO cells expressing equine FucT-VII and/or C2GnT. P-selectin-IgG chimera binding by ePSGL-1 transfected into CHO cells only occurred when both FucT-VII and C2GnT were expressed, establishing that fucosylation and core-2 branching are required as post-translational modifications for high...
Smith ML, Chapman CB.An acepromazine (ACP) hapten was synthesised, coupled to bovine serum albumin and injected into a horse to produce antibodies to the drug. A competitive ELISA was developed whereby ACP attached to the solid phase via lysozyme competed with free ACP present in phosphate buffered saline, horse serum or horse urine for limiting amounts of antibody. The assay could detect the presence of ACP and, or, some of its metabolites in horse urine for at least 25 hours after intravenous injection of 0.1 mg kg-1 ACP maleate, but because of non-specific interference, horse serum could not be used. As little ...
Magnarelli LA, Ijdo JW, Van Andel AE, Wu C, Oliver JH, Fikrig E.To test serum samples of dogs and horses by use of class-specific recombinant-based ELISA for establishing a diagnosis of granulocytic ehrlichiosis attributable to infection with organisms from the Ehrlichia phagocytophila genogroup. Methods: Serum samples from 43 client-owned dogs and 131 horses (81 with signs of acute illness and 50 without signs of disease). Methods: Serum samples were analyzed, using ELISA with a recombinant 44-kd protein antigen for IgM and IgG antibodies to the human granulocytic ehrlichiosis (HGE) agent (NCH-1 strain). Western blot analyses, using infected human promyel...
Horká M, Kubíček O, Kubesová A, Rosenbergová K, Kubíčková Z, Šlais K.Influenza A is viral disease, which is a cause of yearly epidemics and, potentially, pandemics. The conventional techniques used today are equipment-demanding, time-consuming and laborious. Recently, we have confirmed that the capillary isoelectric focusing is a suitable fast alternative for the verifying of virus purity. In the wide pH gradient of pH range 2.0-7.5 the isoelectric points for subtypes of equine (H3N8) and swine (H1N2) influenza A viruses were determined approximately as 6.6 and 6.5, respectively. In this contribution we have verified these findings using different isolates of d...
Gaĭdamovich SIa, Pomelova VG, Lavrova NA, Mel'nikova EE, Sokolova MV, Kharitonenkov IG, Zlobin VN.Potentialities of differentiation between Venezuelan equine encephalomyelitis (VEE) complex viruses by time-resolved fluoroimmunoassay and enzyme immunoassay were studied. For this, 4 test systems were used based on different combinations of native and labeled polyclonal antibodies to VEE virus, strain Trinidad, and monoclonal (MCA) antibody MAK 14-7 to protein EL of this virus. The maximal sensitivity and specificity was achieved in the test system formed from native MCA MAK 14-7 for sensitization of the solid phase and labeled polyclonal immunoglobulins for demonstration of the test results....
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...
