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.
Korkeala H, Stabel-Taucher R, Pekkanen TJ.When 33 horse kidneys were tested for the presence of inhibitory substances by the Bacillus subtilis BGA method at pH 8 and the Micrococcus luteus ATCC 9341 method, 24 were positive and 9 negative. The pH of the seeded M. luteus test medium changed from pH 6.6 before incubation to 8.7 after 24 hours incubation at 30 degrees C. When the same 33 kidneys were tested by the B. subtilis BGA method, medium pH 6, and 15 of them also by the M. luteus method using a medium buffered to pH 6, all were negative. The cadmium concentration of the 33 horse kidneys was found to be 70.17 +/- 81.28 mg/kg wet we...
Targowski SP.The present study describes a two step technique for the separation of mononuclear leukocytes or mononuclear and polymorphonuclear leukocytes from whole equine blood. First, the leukocyte rich plasma was obtained by sedimentation of erythrocytes in the undiluted blood. Subsequently, separation of the different populations of white blood cells was performed by centrifugation with different gradients overlaid with the leukocyte rich plasma. The optimal separation of the mononuclear cells was obtained by the centrifugation of the leukocyte rich plasma overlaying the gradient containing 24 parts o...
Johnson PE, Maister SG, Knowles JR.Phosphoglycerate kinase has been isolated in crystalline form from horse muscle. A convenient isolation procedure is described that yields homogeneous enzyme of specific activity 700 units/mg (30 degrees C). The enzyme is monomeric, and has a molecular weight 47 000. Of the eight cysteine residues in the protein, two react rapidly with Nbs21 with the concomitant loss of the catalytic activity. Since the isolation of phosphoglycerate kinase from yeast (Bücher, 1955) there have been several reports of purification methods yielding enzyme approaching molecular homogeneity, from rabbit muscle (Be...
Scherer WF, Anderson K, Pancake BA, Dickerman RW, Ordonez JV.Seventy-four strains of Venezuelan encephalitis (VE) virus recovered from sentinel hamsters or mosquitoes at enzootic habitats in Guatemala in the two years following the 1969 epidemic-equine epizootic were examined for ability to produce small plaques in Vero African green monkey kidney cell cultures, like isolates obtained during the epizootic. (a) One strain recovered from a sentinel hamster in late October 1969 at an enzootic habitat near the epicenter of the hemagglutination-inhibition (HI) and equine-virulence properties like epizootic virus; this strain retained its small plaque charact...
Poskus E, Zakin MM, Fernámdez HN, Paladini AC.Peptide fragments, obtained from equine growth hormone by cyanogen bromide cleavage and further chemical treatment, were isolated and identified. Their immunological reactivities were tested by hemagglutination and complement fixation methods using rabbit antisera against native hormone. Antigenic determinants were detected in the fragments comprising amino acid sequences 5-72 and 73-123, this last one being predominant. Fragment 124-178 had very low reactivity. Nitration of peptide 73-123 did not modify its immunological properties,but oxidation diminished them. Comparison of the antigenicity...
Ronda GJ, Gaastra W, Beintema JJ.The kinetic parameters Km, k+2 and k+2/Km of the pancreatic ribonucleases (EC 3.1.4.22) from cow, giraffe, horse, rat and lesser rorqual have been determined, using 2',3'-cyclic cytidine monophosphate and 2',3'-cuclic uridine monophosphate as substrates. No large differences were found between the activities of the five enzymes. The relative differences between the activities of the five enzymes are mainly due to differences in the rates of hydrolysis and not to differences in the affinities for the substrates.
Kotts C, Jenness R.Kappa-Casein-like proteins were isolated from the milks of cow, goat, reindeer, horse, rat, and rabbit. When treated with rennin, all of the isolated kappa-casein components yielded para-kappa-casein-like bands on gel electrophoresis. The rate of cleavage of these components with rennin was determined by measuring material soluble in trichloroacetic acid (macropeptide). The curves were characteristic of a limited, specific attack by rennin on these proteins. The goat and reindeer kappa-caseins were nearly as bovine kappa-casein, but the cleavage of horse, rat, and rabbit kappa-casein-like comp...
Marsh JA, Hallett FR, Owen RR.A comparison of methods of preparing the hyaluronic acid of equine synovial fluid for quantitative spectrophotographic analysis is presented. A new method is proposed which appears superior to the previous methods.
Groman EV, Schultz RM, Engel LL, Orr JC.In the reduction of 17beta-hydroxy-5alpha-androstan-3-one to the 3beta-alcohol, horse liver alcohol dehydrogenase utilizes the 4-pro-R hydrogen of NADH whereas the 3(17)beta-hydroxysteroid dehydrogenase of Pseudomonas testosteroni utulized the 4-pro-S hydrogen. These observations provide an exception to the rule proposed by Alworth and Bentley that with regard to the paired methylene hydrogens at C-4 of NADH and NADPH "the stereospecificity of a particular reaction is fixed and does not vary with the source of the enzyme preparation". It is also apparent that for these two enzymes, the selecti...
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.
Stirk SA.A 5 year old Thoroughbred stallion with diarrhoea of unknown aetiology was referred to Davis. Treatment was aimed at terminating diarrhoea and restoring normal fluid status. Laboratory aids were utilised to establish where inbalance and deficits were present. Antibiotics and corticosteroids were used as an adjunct to fluid therapy. The case history and rationale of treatment of fluid disorders resulting from diarrhoea are discussed.
Sogin DC, Plapp BV.Diazonium-1H-tetrazole was tested as a potential active-site-directed reagent for amino acid residues involved in catalysis by alcohol dehydrogenase. In a novel reaction with a protein, diazonium-1H-tetrazole inactivated the enzyme selectively, and almost stoichiometrically, but reacting with the sulfur of a cysteine residue, Cys-174. As a model compound, the tetrazole adduct of free cysteine was prepared. Elementary and spectral analyses of the adduct were consistent with the structure 5-tetrazoleazo-S-cysteine. The adduct absorbs light with a maximun at 316 nm, and is destroyed by irradiatio...
Anderson MG.The distribution of lactic dehydrogenase, aldolase and creatine kinase in various horse tissues was determined. Using polyacrylamide gel electrophoresis the lactic dehydrogenase and creatine kinase isoenzyme composition of horse serum, taken before and after exercise, was studied. Horse tissue isoenzyme patterns were also obtained. By comparing tissue and serum patterns, skeletal muscle was found to be the tissue of origin of the increase in serum lactic dehydrogenase and creatine kinase observed after exercise.
Hafs HD.Often in developing hormone assays, hormones that may interfere with the assay by cross-reaction are not available for testing the validity of the assay. For example, horse TSH was unavailable to test for cross-reaction in an LH radioimmunoassay (RIA). The authors devised an indirect means of accomplishing the same goal, and the evidence from the indirect test of cross-reaction was at least as persuasive as a direct test might have been. Other examples are given of experiments where extensive effort was devoted to validation of steroid RIA, but there were substantial quantitative differences i...
Dubin A, Koj A, Chudzik J.Cytoplasmic granules were isolated from horse blood polymorphonuclear leucocytes by the heparin method and extracted with 0.9% NaCl by repeated freezing. Soluble proteins were separated on a column of Sephadex G-75 followed by chromatography on a column of CM-Sephadex with a NaCl gradient. Gel filtration, density-gradient centrifugation, isoelectric focusing and 0.1% sodium dodecyl sulphate/polyacrylamide-gel electrophoresis at pH 7.0 and at pH 4.5 were used to determine molecular parameters of proteinases. Three enzymes hydrolysing both casein and N-benzyloxycarbonyl-L-alanine nitrophenyl est...
Aberle ED, Judge MD, Kirkham WW, Page EH, Crawford BH.Frozen sections of equine musculus semitendinosus were examined for myosin adenosine triphosphatase (ATPase) and reduced nicotinamide adenine dinucleotide-tetrazolium reductase (NADH-TR), using standard histochemical procedures, and the proportions of the various fiber types and average fiber sectional size were determined. With ATPase staining, approximately 70% of the fibers were classified as alpha fibers (ATPase positive), and 30%, as beta fibers (ATPase negative). In addition, 2 populations of alpha fibers could be readily distinguished on the basis of the intensity of the ATPase reaction...
Seybert DW, Moffat K, Gibson QH.The reactions of horse globin reconstituted with proto-, deutero-, and mesoheme have been examined by equilibrium and kinetic methods. In virtually all reactions studied, mesohemoglobin displays the more extreme functional behavior, whereas deuterohemoglobin exhibits behavior which is either very similar to native hemoglobin or intermediate between the two. Our kinetic and equilibrium results indicate that the primary effect of heme modification on the functional properties of hemoglobin is to alter the intrinsic reactivities of the deoxy and liganded conformations. Heme modification does not,...
Hart LT, Braymer HD, Larson AD, Broussard EA.Equine infectious anemia (EIA) antigen extracted from the spleen of horses infected with EIA virus was purified by pH treatment, (NH4)2SO4 fractionation and affinity chromatography. The homogeneity of the antigen was indicated by sedimentation rate and sedimentation equilibrium experiments. A S20,w of 0.51 was determined and a molecular weight of 7600 was calculated from sedimentation equilibrium analysis. The amino acid composition of the pure antigen indicated the antigen is an acidic protein. Employing radical immunodiffusion (RID) and pure antigen a method for quantitating antigen content ...
Løowenstein H, Markussen B, Weeke B.Three major allergens of horse hair and dandruff have been isolated. The fractionation procedures involved various combinations, described in detail, of ethanol precipitation below --5degreesC, cation- and anion-exchange chromatography, and gel filtration. UV absorption, quantitative immunoelectrophoresis and RAST inhibition were used to monitor the separations. Protein impurities constituted less than 5% in all cases. The molecular weights of the isolated proteins were 1.9 X 10(4), 5.1 X 10(4) and 3.1 X 10(4) daltons, respectively. The pIs were determined as 4.1, 3.8 and 3.9, respectively. Th...
Tepliakova TV, Efremova EA, Riabchikova EI.The carnivorous fungi hyphomycetes are natural enemies of soil nematodes. Laboratory tests examining the effect of the effective strain Duddingtonia flagrans T-89 on equine strongyle larvae have indicated that their size can be reduced 5-48-fold under the action of the fungus. Using helminth-infected mice as an example has ascertained that when the animals are fed a biopreparation, the chlamydial spores of the carnivorous fungus D. flagrans remain viable and continue their development in the excrements. The dead nematodes show cell structural impairments in all tissues and organs, which may be...
Märki U, Osterhoff DR.A method using methotrexate for horse lymphocyte cell synchronization and thymidine for incorporation into DNA replication is described. This method provides a powerful technique for the study of chromosomal abnormalities and detailed analysis of chromosomal replication patterns. The determination of horse karyotypes with many similar chromosomes needs a special method which reveals the numerous and informative stages of the cell cycle. Horse lymphocyte cell cultures treated with colcemid (20 min) and harvested 6 hours after the release of the 17 hour-block with methotrexate show the best resu...
Ollis GW, Morin LA, Visser AL.Through collaborations with Alberta Health and Wellness; Alberta Agriculture, Food and Rural Development; Alberta Sustainable Resource Development; and Alberta Environment, a surveillance program was implemented to detect West Nile virus (WNV) in Alberta and to explore the distribution of the virus in mosquito pools, wild birds, humans, and horses. The surveillance in horses was to detect the presence of the virus in Alberta and to explore possibilities of reducing the risk of infection in both humans and horses. This report gives the frequency, distribution, clinical symptoms, and associated ...
Gummow B, Herr S, Brett OL.A complement fixation test, using round-bottomed microtitration plates and an 8 channel microdiluter, based on that used for brucellosis by Herr, Huchzermeyer, Te Brugge, Williamson, Roos & Schiele, 1985, has been developed for use on the sera of horses to detect antibodies to the contagious equine metritis organism. The results with 2 known positive sera tested 116 times in 27 separate tests were reproducible for the most part within a twofold range. They seldom exceeded these limits and never exceeded a fourfold range. The test itself is capable of being carried out within 90 min. The test w...
Babel I, Stella RC, Prado ES.Previous experiments indicated that horse urinary kallikrein (UK) hydrolyzes salminei- e and polyarginine, a but not polylysine. This paper reports the action of UK on bradykinyl-serine, methionyllysyl-bradykinin and lysyllysyl-bradykinin.
Ricketts SW, Rossdale PD.The authors discuss the value of a practice laboratory to the equine clinician and its priorities. Laboratory examinations of particular value are described in relation to their clinical application. The need to establish normal values according to laboratory and horse population is stressed. Tables of normal parameters related to age groups of horses in the authors' practice are presented.
Bondesson U, Johansson IM.This study demonstrates the development of a method using gas chromatography-mass spectrometry for determining nefopam, a non-narcotic pain reliever that is sometimes abused in horse doping, in equine plasma. Background […]
Dumasia MC, Houghton E.The research investigates the biosynthesis of a particular isomer called 5(10)-estrene-3 alpha, 17 beta-diol in stallion testes and how it affects the formation of 19-nor steroids and oestrogens. Summary of […]
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.
Marqués FJ, Higgins S, Chapuis R, Waldner C.Circulating l-lactate concentration is commonly measured in hospitalized horses by sampling from indwelling intravenous (IV) catheters. However, there are no published evidence-based recommendations to prevent contamination by lactated Ringer's solution (LRS). Objective: Withdrawing 10 mL of blood from the LRS-containing extension set connected to the IV catheter before obtaining the sample for analysis should be adequate to obtain accurate measurement of blood lactate concentration (BLC). Methods: Thirty-three adult hospitalized horses receiving constant rate infusion of LRS. Methods: Immedia...
Nishita T, Deutsch HF.Equine muscle carbonic anhydrase (CA-III) behaves like ubiquitin in undergoing extensive acylation of N epsilon-lysine residues upon reacting with p-nitrophenyl esters. The enzyme undergoes extensive carbamoylation of lysine residues when reacted with carbamoyl phosphate. The modification of from 6 to 7 lysine residues results in the production of a series of more anodic electrophoretic components. The derivatization of the lysine residues leads to a marked decrease in the enzyme's ability to hydrate CO2. The equine CA-III possesses both acid and alkaline phosphatase activities in contrast to ...
DURAN-JORDA F.This research article investigates the significance of eosinophil cells in horses and camels, focusing on the large granules found in these cells and their similarities to smaller red blood cells. […]
Pihl TH, Engelhart KE, Houen G.Polyclonal antibodies are relatively easy to produce and may supplement monoclonal antibodies for some applications or even have some advantages.The choice of species for production of (peptide) antisera is based on practical considerations, including availability of immunogen (vaccine) and animals. Two major factors govern the production of antisera: the nature of adaptive immune responses, which take place over days/weeks and ethical guidelines for animal welfare.Here, simple procedures for immunization of mice, rabbits, sheep, goats, pigs, horses, and chickens are presented.
Oge S, Oge H, Yildirim A, Kircali F.Necropsies on 43 horses, 35 donkeys and two mules slaughtered in Ankara, Turkey, revealed that 12 (15%) of the equines harboured adult Setaria equina. When blood samples were checked for microfilariae, using Knott's method and a combination of membrane filtration followed by histochemical staining for acid phosphatase (AP), only three (4%) of the animals were found to be microfilaraemic. When stained for AP, the S. equina microfilariae exhibited diffuse red staining over the entire body, including the sheath, with brighter staining around the anal and excretory pores. Application of Knott's me...
Finno CJ.In the field of equine muscle disorders, many conditions have a genetic basis. Therefore, genetic testing is an important part of the diagnostic evaluation. Validated genetic tests are currently available for 5 equine muscle disorders: hyperkalemic periodic paralysis, malignant hyperthermia, glycogen branching enzyme disease, type 1 polysaccharide storage myopathy, and myosin heavy chain myopathy. These diseases should be tested for in the appropriate breeds with clinical signs of disease or as part of breeding management. Genetic testing in veterinary medicine is not regulated, and therefore,...
Fujimoto R, Kuchida M, Ban-Tokuda T, Matsui H.The equine large intestine harbors a diverse array of symbiotic microorganisms. Disruptions in the gut microbiota can lead to various diseases in horses. Probiotics offer promising avenues for enhancing equine health and performance. However, commercial formulations lack robust scientific validation. This study aimed to isolate and identify Lactobacillaceae and Bifidobacterium spp. from horse feces to explore their potential as probiotics. Fecal samples from Thoroughbred horses were subjected to isolation procedures. Lactic acid-producing bacteria were isolated using specific media and identif...
Hidalgo M, Ortiz I.Sperm vitrification is a novel-assisted reproductive technique that is increasingly gaining relevance in the last years. This technique allows to cryopreserve sperm from valuable stallions and donkeys without the exposure to permeable cryoprotectants, particularly toxic for the gametes of these species.This chapter aims to describe the current range of methodologies available that are key to ensure sperm quality after vitrification and warming of stallion and donkey sperm.
Miró J, Martínez-Rodero I, Yeste M, Catalán J.Cryopreservation is currently the only strategy for long-term conservation of equine sperm. To get optimal post-thaw sperm survival, carefully following each step of the freezing protocol is crucial. First, one needs to obtain and exhaustively analyze an ejaculate of good sperm quality. Then, the seminal plasma is removed by centrifugation, and the resulting pellet is resuspended in a certain volume of the freezing medium to reach the right sperm concentration. Finally, sperm samples are packaged into 0.5-mL straws, cooled, and frozen using an automatic, controlled-rate freezer. Once the tempe...
Lab on a chipMay 28, 2025
Volume 25, Issue 11 2795-2796 doi: 10.1039/d5lc90048d
Heidenberger J, Reihs EI, Strauss J, Frauenlob M, Gültekin S, Gerner I, Toegel S, Ertl P, Windhager R, Jenner F, Rothbauer M.Correction for 'The effect of cyclic fluid perfusion on the proinflammatory tissue environment in osteoarthritis using equine joint-on-a-chip models' by Johannes Heidenberger et al., Lab Chip, 2025, 25, 2256-2269, https://doi.org/10.1039/d4lc01078g.
Faulk C.Sequencing a genome by students has now become practical as we demonstrated with our recent publication of the Przewalski's horse (Equus ferus przewalskii) genome. In this review, I describe my experience teaching genome assembly in the classroom. In my course, students sequenced, assembled, and published a high-quality genome for Przewalski's horse using Oxford Nanopore long-read sequencing with only $4000 of materials. Along with the genome, we assembled the mitochondrial genome, sequence variants, predicted gene annotations, and DNA methylation levels. Our genome statistics far exceeded the...
Dersch K, Kreuder A, Wang C, Troy J.Surgical site infections (SSI) are a common postoperative complication of equine orthopedic surgery and often increase morbidity and mortality. Objective: This study aimed to determine if covering surgical instruments during surgery decreased bacterial environmental air contamination during early opening of surgical packs (EOSP). The authors hypothesized that covering instruments with sterile surgical towels would reduce the total number of bacterial colonies to which the instruments were exposed. Methods: Blood agar plates (BAP) were allocated into two groups (non-covered vs. covered with a s...
Aboelmaaty AM, El-Seadawy IE, Kotp MS, Mohamed AA, El-Debaky HA, El-Badry DA, Anwer AM, Tsvetkov T, Daneva T, Elgabry MA.Alpha-tocopherol is one of the non-enzymatic lipophilic antioxidants. Butylated hydroxytoluene (BHT) is a synthetic analog with similar modes of action in protecting the cryopreserved sperms. Objective: This study hypothesized that a certain concentration of any antioxidant is suitable for improving the post-thaw semen quality of stallions. Methods: To determine the optimum BHT concentration, a synthetic antioxidant similar to vitamin E in potency and scavenging oxidative stress power in concentrations of 0.0, 0.25, 0.50, 1.0, 2.0, and 4.0 mM/ml were added to semen extender. The post-thaw sper...
Wong KS, Cheung HW, Choi YC, To NS, Wan TSM, Ho ENM.Recombinant human follistatin (rhFST) is a potential performance-enhancing agent owing to its stimulating effect on muscle growth. Administration of rhFST to athletes is prohibited in human sports by the World Anti-Doping Agency (WADA) and in horseracing according to Article 6 of the International Agreement on Breeding, Racing and Wagering published by the International Federation of Horseracing Authorities (IFHA). For effective control of the potential misuse of rhFST in flat racing, methods for screening and confirmatory analysis are required. This paper describes the development and validat...
Ribeiro NG, Silva PD, de Lima Paz PJ, Arabe Filho MF, Listoni FP, Listoni EP, Panegossi LC, Ribeiro MG.Rhodococcus equi is an opportunistic soil-borne bacterium that is eliminated in feces of multi-host animals. An increase in multidrug-resistant R. equi isolates has been reported in humans and domestic animals, and it has been hypothesized that the treatment of R. equi in foals could increase the selective pressure on multidrug-resistant isolates and favor human infections by resistant isolates. We investigated the in vitro antimicrobial susceptibility/resistance of 41 R. equi strains from humans, which were isolated from patients with pulmonary signs, using 19 antimicrobials from 10 distinct ...
Ellison RS, Jacobs RM.The main purpose of this study was to ascertain whether isoelectric point determination of alkaline phosphatase (AP) using an isoelectric focusing technique on agarose gels could define the isoenzymes present in healthy equine serum. The isoelectric points of AP extracted from nine tissues ranged from pH 3.5 to 7.5 with all tissues having multiple bands. There was considerable similarity in band pattern among tissues, with only pancreatic and colostral AP having substantially different isoelectric points from the others. Sera contained thirteen bands with isoelectric points ranging from pH 3.5...
Mancini RA, Suman SP, Konda MK, Ramanathan R, Joseph P, Beach CM.Research focused on determining the fundamental mechanisms by which lactate influences color stability has not considered a direct effect of lactate on myoglobin. Thus, the objective of this study was to use Matrix Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry to examine lactate adduction to myoglobin. Equine oxymyoglobin and equine carboxymyoglobin (0.15mM) were incubated with sodium lactate (200mM) at 4 degrees C, pH 5.6 in 50mM sodium citrate buffer or at 37 degrees C, pH 7.4 in 50mM sodium phosphate buffer, simulating typical meat storage and physiological condition...
Thomas DP, Foley JP.Detection of prohibited substances in equestrian sports typically involves time-consuming and tedious sample-preparation methods. Micellar LC (MLC) allows for direct injection of equine serum to detect prohibited NSAIDs. Results: The method was linear over the range of standards examined, with recoveries of 94.2-95.1% for phenylbutazone (12-18 µg/ml), and 83.9-88.7% and 87.9-105.0% for diclofenac and flunixin, respectively (0.1-1.2 µg/ml). The limit of detection was 0.1 µg/ml for all compounds and the limit of quantitation was 0.2 µg/ml for phenylbutazone and 0.3 µg/ml for diclofenac and ...
Aggarwal BB, Farmer SW, Papkoff H, Stewart F, Allen WR.Serum of the pregnant donkey, like that of the mare, contains a gonadotrophin of chorionic origin. The chorionic gonaditrophin of the donkey (dCG) has been isolated in purified form from the serum of pregnant donkeys using methodology previously employed for the purification of pregnant mare chorionic gonadotrophin (eCG). Unlike eCG, dCG is predominatly an LH in biological tests. In the in-vitro rat Leydig cell assay, dCG was as active as eCG, but in the in-vitro rat seminiferous tubule assay for FSH and in the augmentation assay, dCG was considerably less potent than eCG (1-10%). Specific rat...
