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Topic:Laboratory Methods
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.
Analysis of a complex antigenic site on horse cytochrome c. Of the antigenic determinants so far identified for cytochrome c, only one involves more than a single amino acid substitution between the immunogen and host proteins. Both a threonine at position 89 and a glutamic acid at position 92 control one of the three antigenic sites identified in horse cytochrome c, as expressed in rabbits. Three antibody subpopulations, all directed against this region of the molecule, were isolated from the serum of a single rabbit by adsorption on a series of insolubilized cytochromes c. Antibody fluorescence quenching titrations with a variety of cytochromes c wer...
Activity of adenosine deaminase and purine nucleoside phosphorylase in erythrocytes and lymphocytes of man, horse and cattle. 1. Activities of ADA and PNP were measured in erythrocytes and lymphocytes of man, horse and cattle. 2. In bovine hemolysates both enzyme activities are low when compared with activities in human hemolysates. In horse hemolysates both enzyme activities are virtually absent. 3. Enzyme activities are consistently lower (about 50%) in intact lymphocytes than in sonicated lymphocytes. This finding suggests that the uptake of nucleosides is rate-limiting for both enzymes in intact lymphocytes. 4. The activity of ADA in horse lymphocytes is comparable to that in lymphocytes of patients with severe c...
Demonstration of equine infectious anemia virus in primary leukocyte cultures by electron microscopy. Electron microscopy was used to demonstrate the presence of viral particles in primary cultures of leukocytes taken from a horse after SC inoculation with the Wyoming strain of equine infectious anemia virus. Unlike previous studies, the exposure virus was not passaged through cell culture prior to horse inoculation. Cultures were begun approximately 1 week before and 1 week after the 1st pyrexic period after inoculation. In both samples, viral particles and cytoplasmic alterations were observed resembling those previously reported in equine infectious anemia virus and other retravirus-infecte...
Lactoperoxidase-catalyzed iodination of horse cytochrome c:monoiodotyrosyl 74 cytochrome c. Iodination of horse cytochrome c with the lactoperoxidase-hydrogen peroxide-iodide system results initially in the formation of the monoiodotyrosyl 74 derivative. This singly modified protein was obtained in pure form by ion exchange chromatography and preparative column electrophoresis. It shows an intact 695 nm absorption band, the midpoint potential of the native protein, a nuclear magnetic resonance spectrum which indicates an undisturbed heme crevice structure, a normal reaction with antibodies directed against native horse cytochrome c, and circular dichroic spectra in which the only cha...
A subunit-sized butyrylcholinesterase present in high concentrations in pooled rabbit serum. A butyrylcholinesterase of mol.wt. approx. 83000 was observed in pooled rabbit serum. The enzyme was named monomeric butyrylcholinesterase to distinguish it from the larger oligomeric butyrylcholinesterase of horse and human serum whose subunits are the same size as the monomeric enzyme. The active-site concentration of monomeric butyrylcholinesterase in the pooled serum was 0.18mum, which is five times the concentration of butyrylcholinesterase in pooled horse serum. This was surprising, since the horse serum is regarded as a rich source of butyrylcholinesterase, whereas rabbit serum is not g...
Erythrocyte rosette formation of equine peripheral blood lymphocytes. Erythrocyte rosette (ER) formation of equine peripheral blood lymphocytes (PBL) was characterized. Guinea pig and, to a lesser extent, human erythrocytes formed ER; cat, cow, dog, hamster, mouse, rat, and sheep erythrocytes showed negligible rosetting properties. Conditions of the assay were varied to determine which procedure allowed the largest percentage of rosette formation. The PBL from 20 normal horses were then assayed, averaging 38 +/- 2% ER. To characterize the erythrocyte receptor as being on T or B cells, equine thymocytes from 6 foals were assayed; the thymocytes formed an average ...
Studies related to the metabolism of anabolic steroids in the horse: 19-nortestosterone. 1. The metabolism of 19-nortestosterone in a cross-bred horse has been studied using 14C-labelled material. 2. Two neutral metabolites isolated from urinary extracts by column chromatography were identified as isomers of 3-hydroxyestran-17-one and estrane-3,17-diol by g.l.c.-mass spectrometry. 3. The stereochemistry of the two metabolites has been investigated by comparison of the retention times of their trimethylsilyl derivatives with those of standard steroids of known configuration.
Staining of glycosaminoglycans in intervertebral disc cells. Disc material from horse, ox, sheep, pig, dog and cat was stained by the Alcian-blue-critical electrolyte concentration technique and with the standard and two-step periodic acid Schiff methods. The effects of pretreatment with hyaluronidase and with chondroitinase was also evaluated. There appears to be a small increase in total cellular glycosaminoglycan content with age in all species: cellular material of high molecular weight however only increases in aged animals. The degree of sulphation of cellular glycosaminoglycans does not vary with age or with position in the disc.
Circular dichroism of porcine, bovine, and equine pancreatic phospholipases A2 and their zymogens. Unusual conformations simulating helix content. Conformation of porcine, bovine, and equine pancreatic phospholipases A2 (EC 3.1.1.4) and their zymogens was studied by the circular dichroism (CD) probe in the far and near ultraviolet spectral zones.
All these phospholipases and their zymogens displayed CD curves suggesting the presence of moderate amounts of α-helical conformation. However, on the basis of known primary structure and recent X-ray structural analysis of prophospholipase A2 crystals (Drenth, J., Enzing, C.M., Kalk, K.H. and Vessies, J.C.A. (1976) Nature 264, 373–377), it has to be concluded that the positive CD band cen...
Studies on a number of erythrocytic enzymes and intermediate products of equine erythrocyte metabolism. 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...
Nucleolar fragmentation in cells infected with alphaviruses (39886). No abstract available
Semisynthetic cytochrome c. Horse heart cytochrome c can be split with cyanogen bromide into a heme peptide (residues 1-65) and a nonheme peptide (residues 66-104). In a process involving (i) complex formation between the two fragments and (ii) restoration of the severed peptide linkage, a fully active cytochrome c preparation can be re-formed. Use has been made of this process to couple the heme peptide to peptide 66-104 synthesized by the Merrifield solid-phase procedure. The semisynthetic product formed in this manner is indistinguishable from reconstituted cytochrome c prepared with nonsynthetic peptide 66-104.
[Fibrinogen level in clinically healthy horses]. Various sets of horses were examined in view of the necessity of widening the range of biochemical substances for diagnostic purposes in the field of veterinary medicine. The aim of the investigations was to obtain basic information on fibrinogen level and its variability. The average value stated was 280 mg% s = 89, sx = 12, V% = 32). The fibrinogen level in horses of the pronounced oxidation type was lower than in those with reduced metabolism, the difference being connected with the speed of sedimentation of erythrocytes. Repeated examination of a part of the set of horses (n = 10) on three...
Methods in the evaluation of antiparasitic drugs in the horse. The critical test is the primary method used for the efficacy evaluation of drugs against the major internal parasites (bots, ascarids, large strongyles, small strongyles, and pinworms) of the horse. The critical test determines: (1) spectrum of activity, (2) effectiveness of removal, (3) pattern of discharge, and (4) physical condition of each species of these parasites. General characteristics of the major parasitisms of the horse are discussed briefly. Criteria of the critical test also are considered including: (1) number of tests, (2) strain variation and drug resistance, (3) selection of...
Chromatographic separations of alphavirus strains by hydroxylapatite. Hydroxylapatite column chromatography methods were developed to characterize selected alphavirus populations. Different conditions of pH and phosphate molarity were required to obtain satisfactory elution profiles and separations for Western equine encephalomyelitis virus strains, compared with Eastern equine encephalomyelitis virus and Semliki Forest virus strains. Raising the pH of the buffers effected earlier elutions of all viruses. Selection of phosphate gradients with more gentle slopes and adjustment to the proper pH effected better separations of virus subpopulations. Elution profiles ...
Effects of storage on the methaemoglobin content of equine blood. Equine blood containing different levels of methaemoglobin was stored under varying conditions and the methaemoglobin content was monitored during the storage period. Only under aerobic storage at 4 degrees C did the methaemoglobin content of all samples appear to remain stable.
Practical methods of determining serum immunoglobulin M and immunoglobulin G concentrations in foals. Serum concentrations of immunoglobulin M (IgM) and immunoglobulin G (IgG) can be determined in the horse with a satisfactory degree of accuracy, using commercially available reagents. Selected lots of anti-human IgM can be used in precipitation tests to detect and quantitate equine IgM. Commercially available anti-equine IgG tended to overestimate the amount of IgG in single radial immunodiffusion tests. Even with these limitations, commercial reagents can be used to differentiate immunodeficiency disorders of foals, including combined immunodeficiency and failure of passive transfer of colost...
Inhibition of the growth of some strains of Mycoplasma mycoides subsp mycoides by the blood of certain horses. 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...
The oxidation of ferrocytochrome c in nonbinding buffer. The apparent equilibrium constant and rate of oxidation was investigated for the reaction of cytochrome c with iron hexacyanide. It was found that if horse heart ferricytochrome c was exposed to ferricyanide (to oxidize traces of reduced protein) the cytochrome subsequently, even after extensive dialysis, had an apparent equilibrium constant different from that of electrodialyzed protein. The effect of ferricyanide ion apparently cannot be removed by ordinary dialysis. The ionic strength dependence of the apparent equilibrium constant and bimolecular oxidation rate constant was measured in the...
Studies on the antigenicity of an inactivated, aluminum hydroxide adjuvant equine influenza vaccine. An inactivated, aluminum hydroxide adjuvant equine influenza vaccine was tested in horses and guinea pigs to determine the levels of antigen that would elicit maximum serological responses. Vaccine containing serial twofold increments of A/Equi-1/Prague and A/Equi-2/Miami strains of equine influenza virus was administered to random groupings of both types of test animals. The hemagglutination inhibition antibody response for each group was then measured. Results in horses and guinea pigs were compared to determine if the equine serological values could be related to a potency test in laborator...
Lactic acid concentration in peritoneal fluid of normal and diseased horses. Peritoneal fluid from each of 15 clinically healthy horses and five horses with acute abdominal disease was evaluated for lactic acid concentration. The normal range was 2-7--13-4 mg/dl. Simultaneous blood and peritoneal fluid samples from healthy horses revealed consistently lower lactic acid concentrations in the peritoneal fluid than in the blood, whereas peritoneal fluid lactic acid levels were consistently greater than blood levels in the diseased horses. The diseased horses had highly significant (P less than 0-005) increases in both blood and peritoneal fluid lactic acid concentrations ...
Direct radioimmunoassay of progesterone in mare plasma. A rapid and low cost radioimmunologic procedure for progesterone assay in mare plasma is proposed. Radioimmunoassay is performed directly on 10 microliter of unextracted plasma. Free progesterone is adsorbed on dextran-charcoal, then the aqueous phase is decanted and extracted by 1 ml of scintillation fluid. Counting is performed directly on this two-phase system. Results are comparable to those obtained with radioimmunoassays using extracted plasma.
