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.
Miquelis R, Simon C.1. The subcellular particles of horse and rat thyroids were fractionated in a B XIV zonal rotor on a non-linear gradient of Ficoll after labelling with radioactive iodine in vitro (horse) or in vivo (rat). In the horse, the resulting fractions were analysed for radioactive iodine, protein and enzymes representative of certain subcellular particles. In the rat, iodine turnover and thyrotrophin stimulation were studied. 2. The population of iodinated particles could be subdivided into three main classes, characterized by differences in beta-galactosidase and acid phosphatase content and position...
Carter EI, Valli VE, McSherry BJ.The peripheral blood cells from Standard bred horses were subjected to procedures which will separate equine peripheral blood cells with good precision and efficiency into red cell, leukocyte, and platelet fractions. The separated cells have normal morphology and the differential count of the separated granulocytes and lymphocytes is unchanged from that of the original sample.
Roberts MC.SUMMARY
A D(+) xylose absorption test has been standardised for use in the horse. A dose of 2 g. xylose/Kg. body-weight is required to produce a maximum plasma xylose level of 30 mg./100ml. after two hours. There are limitations to the procedure being used as a routine clinical screening test of small intestinal dysfunction in the horse.
RÉSUMÉ
On a standardisé un test d'absorption du D(+) xylose pour le cheval.
Une dose de 2 gr de xylose par kilo de poids vif est nécessaire pour produire une élevation du xylose plasmatique jusqu'à 30 mg par 100 ml après deux heures.
Il y a d...
Kadivar A, Rashidzadeh H, Davoodian N, Nazari H, Dehghani Tafti R, Heidari Khoei H, Seidi Samani H, Modaresi J, Ahmadi E.Accurate diagnosis of foetal sex in pregnant mare is helpful for many breeders, both for private or commercial purposes. In this study, in order to pre-natal foetal sexing in equine, we used TaqMan duplex real-time PCR to detect the specific regions of SRY and TSPY genes on extracted cell-free foetal DNA from maternal blood. Peripheral blood samples from 50 pregnant Arabian mares with singleton foetuses were collected. Cell-free foetal DNA was extracted from maternal plasma, and duplex real-time PCR assays were performed with TaqMan probes and primers. Amplification of glyceraldehyde-3-phospha...
Dusek J.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...
Fatemikia H, Kamyab M, Movahed A, Sadeghi M, Kim E, Behdani M, Mohammadpour Dounighi N, Shahrivar M, Seyedian R.In this study, the neutralizing abilities of the equine and the recently introduced camelid antivenoms on the hemodynamic parameters (inotropism, chronotropism, and arrhythmogenicity) were assessed following envenomation by venom in rats. Methods: At first, the electrophoretic profiles of both products were obtained by using the SDS-PAGE method (12.5%) and stained with Coomassie blue and silver nitrate. Secondly, different doses of the camelid antivenom (10, 50, and 100 µl) were given intravenously in 10 min before venom injection (400 µg/rat). The neutralizing potencies of camelid and equi...
Fukushima JG, Cascone O, Santomé JA, Biscoglio de Jimenez Bonino MJ.Reactivity of histidine residues in equine growth hormone to ethoxyformic anhydride was studied. The existence of two kinetically different sets was demonstrated: one of them including only the slow reacting histidine 169 (k = 0.164 min-1) and the other containing fast reacting histidines 19 and 21 (k = 0.892 min-1). A correlation between the decrease in the capacity to compete with 125I-labeled hormone for rat liver binding sites and the degree of ethoxyformylation of the fast group was found. Circular dichroism studies indicated no significant conformational changes in the protein with all t...
Campbell-Thompson M.Equine oxyntic mucosal cells were obtained by sequential exposure to pronase and collagenase. Acid production by parietal cells was assessed by uptake of [14C]aminopyrine (AP), a weak base that accumulates in intracellular acidic spaces. Incubation for various times revealed a maximal AP uptake at 10 minutes for histamine and carbachol. Similar secretagogue responses were observed for parietal cells from the mucosal cell preparation or after enrichment by elutriation. Histamine and isobutyl-methylxanthine (IBMX) stimulated AP uptake with a dose-dependent response and maximal effective concentr...
Netherwood T, Chanter N, Mumford JA.There have been several case reports of foal diarrhoea associated with Clostridium perfringens. However, there has been no epidemiological assessment of the strength of the association of these bacteria with foal diarrhoea or of their relative importance. To prepare methods for such a study, the success of different cultural techniques for the isolation of C perfringens was examined with respect to the various physiological states of the bacteria. The germination and growth of C perfringens NCTC 8239 endospores of differing maturity were favoured by different pre-treatments which failed to rec...
Roberts CJ, Jackson LS.The development, validation, and application of an ELISA for dexamethasone in equine urine is described. The drug-protein conjugate was immobilised in microtitre plate wells and antiserum raised against the same drug-protein conjugate was allowed to compete with sample or standard drug and the immobilised drug-protein conjugate. The proportion of antiserum binding to the immobilised drug-protein conjugate was detected using a biotinylated protein G/extravidin-alkaline phosphatase complex in situ and measurement of the substrate product. The method was used to detect the presence of drug-derive...
Pellegrini A, Zweifel HR, von Fellenberg R.Several publications have described in the past properties of partly purified horse alpha 2-macroglobulin (alpha 2M) which are strikingly different from the human alpha 2M. Horse alpha 2M was therefore isolated to purity by classical procedures, i.e. affinity chromatography, ion exchange chromatography and gel filtration, and its properties are compared with those of its human counterpart. The molecular weight of the native protein and its subunits, the isoelectrofocusing pattern and the change in electrophoretic mobility caused by interaction with protease were similar to those of human alpha...
Weiser MG.Erythrocyte volume distribution curves (erythrograms) were determined on a total of 300 blood samples from healthy dogs, cats, horses, and cattle (dairy cows). An index of anisocytosis was determined for these animals. Erythrograms were highly reproducible, and the mean corpuscular volumes determined from erythrograms compared well with those determined from hemograms. Bovine and equine erythrocyte volumes were found to be stable after the blood was stored at 4 C for 24 hours. Under the same conditions, canine and feline erythrocytes increased slightly in volume. After incubation of blood dilu...
Lindner A, Will Y, Wilkniss E, Chrispeels J.The influence of duration and temperature of storage on hormone levels of whole blood, plasma and serum of horses was investigated. Using FPIAs cortisol, T4 and T-uptake could be measured while the T3-FPIA did not work appropriately. Serum and Plasma stored under the same conditions did not show any difference in cortisol, T4 and T-uptake values. In frozen heparinized plasma samples analysed on different days the cortisol and T4 concentrations fluctuated markedly. The T-uptake values were rather stable. The smallest day by day changes of cortisol and T4 in plasma were found when storing the sa...
Black SB, Stenhouse AM, Hansson RC.This paper details various rapid and sensitive methods for the extraction and derivatisation of propranolol, metoprolol, sotalol, atenolol, pindolol, timolol, oxprenolol, alprenolol and penbutolol in equine urine and in human post mortem whole blood and urine. Three solid-phase extraction methods are described involving the use of either XtrackT XRDAH515, Bond Elut Certify or Sep-Pak C18 cartridges. Two derivatisation methods are also described involving the formation of cyclised silyl or pentafluoropropionate derivatives with either chloromethyldimethylchlorosilane or pentafluoropropionic anh...
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...
Raengsakulrach B, Staczek J.Equine cytomegalovirus (ECMV) contains a linear, double-stranded DNA genome composed of a 146-kbp unique region flanked by a pair of 18-kbp direct repeat (DR) sequences at the termini. Cycloheximide, actinomycin D, and phosphonoacetic acid were applied to infected cell cultures to divide viral transcription into immediate-early (IE), early, and late phases. Eight IE transcripts were identified and mapped to two regions (I and II) of the viral genome. Two of these IE RNAs (13.0 and 5.5 kb in size) were transcribed from region I, which is located within the DR regions; these IE genes are diploid...
Kim Y, Seo C, Oh S, Kwak J, Jung S, Sin E, Kim H, Ji M, Lee HS, Park HJ, Lee G, Yu J, Kim M, Lee W, Paik MJ.Loxoprofen is a non-steroidal anti-inflammatory drug of the 2-arylpropionic acid type, which has used to treat musculoskeletal disorders in the horse racing industry. However, it has also used illicitly to mask clinical signs of inflammation and pain in racehorses. Thus, its accurate analysis has become an important issue in horse doping laboratories. In this study, an analytical method of loxoprofen was developed as tert-butyldimethylsilyl (TBDMS) derivative by gas chromatography-mass spectrometry (GC-MS). Characteristic fragment ions of [M-15], [M-57], and [M-139] permitted the accurate and ...
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...
Delbeke FT, Landuyt J, Debackere M.A high-performance liquid chromatographic method to measure plasma and urinary alclofenac levels in equine biofluids is described. Isolation of the drug from plasma is achieved using liquid-liquid extraction with diethyl ether. Reversed-phase C18 solid phase extraction is used for the extraction of free and conjugated alclofenac from urine. The reproducibility and accuracy of the method were well within acceptable limits over the concentration ranges 0-10 and 0-20 micrograms/ml, respectively, for plasma and urine. Starting with 2 ml of plasma, a concentration of 0.1 microgram/ml could easily b...
Greenaway EC, Cunningham FM, Goode NT.Phorbol esters, which activate protein kinase C (PKC), stimulate equine eosinophil superoxide production and adherence. After showing that superoxide production could be inhibited by the nonselective PKC inhibitors, staurosporine and bisindolymaleimide I, the PKC isotypes in equine eosinophils were characterized, because evidence suggests that individual isotypes may play distinct roles in regulating eosinophil function. Western blots demonstrated that equine eosinophils expressed PKC alpha, beta, delta, epsilon, iota, and zeta. However, unlike the equine neutrophil, the majority of the PKC wa...
Hülsemann F, Flenker U, Machnik M, Schänzer W.Sodium bicarbonate administration to horses prior to competition in order to enhance the buffer capacity of the organism is considered as a doping offence. The analysis of the isotopic composition of urinary bicarbonate/CO(2) (TCO(2)) may help to identify an exogenous bicarbonate source, as technical sodium bicarbonate exhibits elevated delta(13)C values compared with urinary total carbon. The isolation of TCO(2) from 60 equine urine samples as BaCO(3) followed by an isotopic analysis shows a significant variability of delta(13)C for TCO(2) of more than 10 per thousand. The delta(13)C of total...
Drbalova J, Musilova P, Kubickova S, Sebestova H, Vahala J, Rubes J.The T cell receptor (TCR) genes (TRA, TRB, TRD and TRG) reside in 3 different chromosomal regions. During the maturation of T lymphocytes, the TCR genes are rearranged by site-specific recombination, a process that also predisposes T cells to aberrant rearrangements. Illegitimate recombination between the TCR genes occurs at a low level in healthy individuals, but this frequency may correlate with the risk of lymphoma. The aim of this work was to investigate interlocus recombination in equids. Illegitimate rearrangements were studied in peripheral blood lymphocytes by FISH with painting and BA...
Knowles EJ, Hyde C, Harris PA, Elliott J, Menzies-Gow NJ.A chemiluminescent immunoassay is commonly employed to measure adrenocorticotrophic hormone (ACTH) concentrations to assist pituitary pars intermedia dysfunction diagnosis. In a previous study, seasonally-dependent assay cross-reactivity to endogenous equine corticotropin-like intermediate lobe peptide (CLIP, ACTH 18-39) was suspected. The present study aimed to demonstrate binding of endogenous equine CLIP to the capture antibody of the ACTH chemiluminescent immunoassay. Liquid chromatography - mass spectrometry (LCMS) methods were optimised to identify selected ions from synthetic human ACTH...
Smith P, Stubley D, Blackmore DJ.Methods were developed for the measurement of superoxide dismutase (SOD), diamine oxidase (DAO) and caeruloplasmin oxidase in the blood of thoroughbred horses. These enzymes were measured in 178 normal thoroughbreds stabled throughout the United Kingdom. The relationships between the activities of SOD, DAO and caeruloplasmin oxidase and the blood concentrations of their associated trace metals (copper, zinc and manganese) were studied in 52 of the thoroughbreds. Trace metals were measured by electrothermal atomic absorption spectrophotometry. No relationships were found between the activities ...
Bird J, Larsen M, Nansen P, Kraglund HO, Grønvold J, Henriksen SA, Wolstrup J.Two sets of dung-derived organisms from soil routinely fertilized with manure (MA) and soil chemically fertilized (CH) were cultured separately in the laboratory. Baermannized organisms from these cultures were added to 20 g of faeces from strongyle-infected horses to form three treatment groups: (i) no soil organisms; (ii) low inoculum of soil organisms containing all organisms present in a suspension of approximately 100 adult female free-living nematodes; and (iii) high inoculum containing those soil organisms present with approximately 1000 adult female free-living nematodes. Three studies...
Gil L, Galindo-Cardiel I, Malo C, González N, Alvarez C.Cholesterol and Equex-STM are frequently added to different commercial and experimental extenders improving postthawing sperm quality. Doses of 125-150 mM of cholesterol from pig liver and 0.5-0.7% of Equex-STM were evaluated in a standard eggyolk extender (Martin et al., 1979). Six ejaculates per stallion from six pure Spanish stallions (6-8 years old) were collected in Martin's extender (B) and different mixtures of 125 mM-0.5% (I), 125 mM-0.7% (II), 150 mM-0.5% (III), and 150 mM-0.7% (IV) were added to original Martin's extender. Samples were frozen in 0.5 mL straws (100 × 10(6...
van Loon JP, de Grauw JC, van Oostrom H.The aim of this study was to compare different methods to determine venous admixture (Q˙/Q˙) in anaesthetized horses. The first objective was to estimate Q˙/Q˙ using jugular venous blood oxygen content (Q˙/Q˙), and a fixed value for the oxygen extraction (F-shunt). The second objective was to assess the influence of blood pressure and positioning on oxygen extraction. The third objective was to perform regression analysis between jugular and mixed venous blood oxygen tensions. Methods: Prospective, experimental trial. Methods: The study was performed with seven warmblood horses that were...
Lee JY, Kang TY, Lee YD, Shin TK.The binding specificities of various lectins, such as the Dolichos biflorus agglutinin (DBA), soybean agglutinin (SBA), and the Bandeiraea simplicifolia BS-1 (Isolectin B4), Triticum vulgaris (WGA), Arachis hypogaea (PNA), and Ulex europaeus (UEA-I) lectins, were studied in the vomeronasal organ of the horse. The microvilli of the vomeronasal sensory epithelium were positive for DBA, SBA, Isolectin B4, WGA, PNA, and UEA-I. The receptor cells showed intense reactivity for DBA and WGA. Lectins were not detected in the supporting cells or basal cells. The Jacobson's glands were positive for WGA a...
Kaneko JJ, Rudolph WG, Wilson DW, Carlson GP.Serum bile acids were fractionated by high-performance liquid chromatography (HPLC) in 13 control and 8 cases of liver disease in horses. The severity and type of liver injury was determined by histopathological examination of biopsy and/or necropsy specimens. The total serum bile acids (tSBA) were determined in these horses by an enzymatic method (SBA-EA) and by summation of the bile acids (SBA-LC) as fractionated by the HPLC. The SBA-LC were generally higher than the SBA-EA in both the controls and liver disease and they did not parallel each other. The primary bile acids, total cholates and...
Bland SA, Blake JW, Ray RS.Mefenamic acid is extracted from biological fluids and is acylated with pentafluoropropionic anhydride to form a derivative possessing high electron affinity. The derivative is analyzed by gas-liquid chromatography with an electron capture detector. The method is particularly valuable for determining drug levels in blood where small sample and/or drug concentrations are available.
Thomas PG, Ball BA.To facilitate the study of interactions between equine spermatozoa and homologous oviduct epithelial cells, we developed an assay to count labelled spermatozoa bound to oviduct epithelial cell (OEC) monolayers and used the assay to compare the binding ability of spermatozoa from different stallions. Washed spermatozoa from three stallions were incubated with the fluorochrome Hoechst 33342 (5 micrograms/ml) for 1 min. Spermatozoa were then layered over confluent monolayers of oviduct epithelial cells in 2 cm2 culture wells. Coculture treatments comprised five concentrations of spermatozoa (10(5...
Monet-Kuntz C, Guillou F, Fontaine I, Combarnous Y.Using a suspension of seminiferous tubule cells, we had previously shown that equine FSH is superactive in the male rat, i.e. that it exhibits a higher biological potency than expected from its binding activity. In this work we investigated equine FSH superactivity in rat, pig and sheep, by comparing in each species the equine FSH with the homologous FSH, both for their binding activities (in a radioreceptor assay using a testicular membrane fraction) and for their in vitro biological potencies (in a plasminogen activator assay using a Sertoli cell-enriched population cultured on plastic). In ...
Spirito S, Campi S, Boquet M, Fernández H, Ferrari M.The aims of this study were to estimate the ability of the Feulgen reaction to identify equine sperm nuclei with different ploidy levels, to determine the frequency of haploid, diploid and polyploid sperm nuclei in the semen of fertile equines and to evaluate the relationship between the nuclear DNA content and the nuclear area. Determination of the ploidy level of Feulgen-stained spermatozoa using a scanning microspectrophotometer was very similar to the subjective estimations made with a light microscope. This indicates that the Feulgen reaction is a simple, inexpensive and reliable techniqu...
