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.
Olitsky PK, Cox HR, Syverton JT.We have studied certain properties, additional to those previously described (3), of the virus of vesicular stomatitis of horses, and of the characteristic biological reactions of the virus of equine encephalomyelitis. It has been found that the virus of stomatitis, ordinarily dermotropic, can acquire neurotropism and the neurotropic encephalomyelitis virus can, in turn, be rendered dermotropic in its action. The neurotropism in both instances is associated with definite, although not pronounced, viscerotropism. Both viruses can bring about a similar infection in the white mouse, rat, guinea p...
Parker JT, McCoy MV.1. The serum of horses immunized with increasing doses of certain anaerobically produced autolysates of pneumococci contain potent neutralizing antibodies for the pneumotoxin. 2. The method for the in vitro titration of these horse antipneumotoxic serums is given.
Abramson HA.1. The cataphoretic velocity of blood platelets (horse) in plasma has been found to be between 40 and 51 micro per sec. per volt per cm. The mean velocity obtained from five horses is . 45 micro per sec. per volt per cm. 2. The cataphoretic velocity of polymorphonuclear leucocytes in similar specimens is practically identical with that of the platelets. This is noteworthy because of the fact that lymphocytes and red cells have different speeds. 3. With spontaneous agglutination of platelets, white cells and red cells, there is no change in the cataphoretic velocity incidental to aggregation. 4...
Friedlander M, Sobotka H, Banzhaf EJ.The precipitin indices for a number of monovalent and polyvalent antipneumococcus sera were determined under known conditions, and found to vary as did the number of protective units. The ratio precipitin index/protective units in monovalent sera was found to lie between 2.8 and 4.8 for Type I and to be about ten times greater for Type III. Lower values were found in polyvalent horses and when mixing heterologous monovalent sera with each other. The influence of the duration of treatment upon the quotient was studied. Several refined and concentrated preparations showed a relative increase in ...
Reimann HA.It is conceivable that a change from the virulent, non-phagocytable S form of Pneumococcus to the avirulent phagocytable R form may take place in pneumococcus disease, but the experiments here reported do not settle the question whether or not this is an important factor in determining the outcome in natural infection. It has been shown experimentally that the degradation from the S form to the R form actually does take place in cultures of Pneumococcus growing in agar subcutaneously embedded in guinea pigs, in agar enclosed in vials subcutaneously embedded in rabbits, and spontaneously in the...
Jungeblut CW.1. A flocculation reaction has been described which occurs between alcoholic extracts of pneumococci and antipneumococcus serum. 2. The reaction appears to be species-specific. It is not strictly type-specific, as slight or moderate cross-reactions occurred between Type I serums and Type II and Type III extracts. 3. The flocculating power of the serum from five horses undergoing immunization with pneumococcus, Type I, did not develop to any extent before the end of the 4th or 5th month. 4. In the case of two of these horses in which it was possible to carry out parallel tests on a larger numbe...
Amoss HL, Marsh P.Experiments were made for the purpose of testing the reaction of protection against infection as a measure of potency of antimeningococcic serum. The results of the experiments were extremely variable and bore no relation to the quality of the sera as determined by the period of immunization of the horses from which they were obtained, or the indications of efficiency based upon their employment in human cases of epidemic meningitis. The results also failed entirely to conform to the agglutination titer of the sera tested and to be affected by the different type forms of the meningococci. We r...
Wadsworth AB, Kirkbride MB.Horses immunized to Type I pneumococci developed serum, 0.1 cc. of which protected against 0.5 cc. of a virulent culture, 0.000001 cc. of which killed mice in less than 40 hours. Protective tests of serum from horses immunized to Type II organisms varied, 0.1 cc. protecting, however, in certain instances against 0.1 and 0.01 cc. of virulent homologous culture. Types I and II sera obtained in our experiments with culture sediment and whole culture did not vary markedly for a given type and corresponded closely in their protective titer with samples of sera received from The Rockefeller Institut...
Hiss PH, Atkinson JP.THE RESULTS OF THE FOREGOING EXPERIMENTS MAY BE BRIEFLY SUMMARIZED AS FOLLOWS: The amount of antitoxic substance obtained by precipitation with magnesium sulphate from the blood-serum of the horse corresponds, as nearly as can be determined by the use of test guinea-pigs, in full to the protective power of the serum from which it is obtained, i. e. the precipitate from 1 cc. of serum will protect against the same amount of toxin as 1 cc. of the serum itself. Equal amounts of the precipitates by magnesium sulphate from immunized and non-immunized horses act differently toward toxin; i. e. the p...
Atkinson JP.1. The globulins of both normal and diphtheria antitoxic serum exhibit chemically toward reagents the same reactions, being precipitated by magnesium sulphate and split up into fractions in precisely the same way. 2. All of the diphtheric antitoxic power of both normal and immunized serum is always carried by the globulin and its fractional precipitates. 3. During the fractional precipitation of the serum globulin of horses immunized from diphtheria toxin and horses not immunized from diphtheria toxin, some of the globulin is lost, likewise at the same time some of the antitoxic power of the g...
The goal of this work was the development of suitable (real-time) RT-PCR techniques for fast and sensitive diagnosis of EAV and for molecular-epidemiological characterisation of viral strains, as an alternative to virus isolation. To this purpose two conventional RT-PCR methods and one real-time RT-PCR were adapted to detect the broadest possible spectrum of viral strains. Several dilutions with Bucyrus strain showed a 100-fold higher sensitivity of real-time RT-PCR and heminested RT-PCR compared to simple RT-PCR. Making use of 11 cell culture supernatants of different EAV isolates and 7 semen...
Shaffer SK, Medjaouri O, Swenson B, Eliason T, Nicolella DP.The ability to quantify equine kinematics is essential for clinical evaluation, research, and performance feedback. However, current methods are challenging to implement. This study presents a motion capture methodology for horses, where three-dimensional, full-body kinematics are calculated without instrumentation on the animal, offering a more scalable and labor-efficient approach when compared with traditional techniques. Kinematic trajectories are calculated from multi-camera video data. First, a neural network identifies skeletal landmarks (markers) in each camera view and the 3D location...
Kirmse L, Thieme K, Doherr MG, Eule JC.To evaluate different laboratory procedures for determining the etiologic diagnosis of equine recurrent uveitis regarding intraocular infection with Leptospira spp. and to establish a diagnostic guideline. Methods: Eighty horses with a history of ERU were ophthalmologically examined. Serum and aqueous humor were collected. Total protein, albumin level, and MAT against Leptospira spp. were evaluated on serum and aqueous humor. PCR for Leptospira spp., EHV-1 and -4 was performed on aqueous humor. Goldmann-Witmer coefficient (GWC) and C-value (CC) were calculated based on MAT. In 42 cases, an add...
So YM, Kwok WH, Yap C, Wong COL, Smalley SGR, Forbes BS, Chow DWY, Ho ENM.This paper describes the detection and longitudinal monitoring of cyclosporine in plasma and urine after subconjunctival implant administration in a horse. Sensitive liquid chromatography tandem mass spectroscopy (LC-MS/MS) methods for detecting cyclosporine in horse plasma and urine have been developed and validated, with estimated limits of detection down to 1 pg/mL in both matrices. The developed methods enabled longitudinal monitoring of cyclosporine levels in blood and urine samples collected over 6 months from a horse that had received an ocular cyclosporine implant.
Friend MM, McGaffigan EM, Hall SM, Staniar WB, Smarsh DN.Equine blood samples are often collected in uncontrolled, distant environments and stored for later analysis. Objective: This study aimed to determine how sample storage methods influence complete blood count (CBC) parameters, hypothesizing that temperature and time elapsed from collection to analysis would affect CBC parameters. Methods: Blood samples were collected from 10 healthy Quarter Horse mares (age: 13 ± 4.5 years old) and stored in EDTA tubes at room temperature (22°C), above ice (8°C), refrigerated (4°C), or in contact with ice (2°C). The IDEXX ProcyteDx was used to obtain CBCs...
Bindi F, Cingottini D, Sorvillo B, Sala G, De Marchi L, Monni G, Celardo S, Bonelli F, Sgorbini M.During the perinatal period, neonates experience a sudden increase in oxygen exposure, predisposing them to oxidative stress. Objective: This prospective, in vivo study aimed to evaluate biomarkers of oxidative stress and antioxidant defence in healthy neonatal foals during the first 48 hours of life. Methods: Ten Standardbred foals were enrolled. Blood samples were collected at birth, 24 hours, and 48 hours after birth and analyzed for biomarkers using spectrophotometry assays. Statistical analysis was performed using non-parametric tests and linear regression. Results: The results revealed s...
Carotenuto SE, Broussard G, Fox GA, Hayes WK.To assess the efficacy of a new equine F[ab'] antivenom (US Department of Agriculture code 6101.05) in removing serum venom and lowering the snakebite severity score (SSS) in animals suffering from naturally occurring viperid envenomation. Methods: Retrospective study, April 2020 to October 2021. Methods: Veterinary hospitals across four North American regions. Methods: Twenty-two dogs, one cat, and one horse. Methods: All patients received the new antivenom intravenously and were assessed for clinical response. Most dogs required one vial for clinical control of signs. Results: Among the dogs...
Pasicka E, Baca M, Popović D, Makowiecki D, Janeczek M.This study presents the sequencing and analysis of mitochondrial DNA (mtDNA) genomes from nine early medieval horse remains excavated across archaeological sites in Silesia region in present day Poland. Methods: Using aDNA extraction protocols optimized for short fragments, combined with target enrichment and high-throughput sequencing, we reconstructed partial mtDNA sequences for seven of the specimens. Results: The authenticity of the aDNA was confirmed through damage pattern analysis. Phylogenetic reconstruction revealed that the specimens belonged to six distinct mtDNA lineages (B, D, E, G...
Ventura Lopes Carvalho B, Neves de Souza MC, Souza Moreira N, Parisi Marliere J, Mesquita Mota JV, Drumond Bento L, de Castro Benitez A, Avanza MFB....Two routes of administration for electrolyte solutions are commonly used in horses with fluid, electrolyte, and acid-base imbalances: intravenous and nasogastric. Despite the frequent use of these routes, there are situations in which they cannot be applied. In such cases, intracecal fluid therapy represents a viable alternative, as this route enables the administration of enteral electrolyte solutions even in animals lacking normal anterior gastrointestinal function. This study aimed to evaluate the effects of a neutral hypotonic enteral electrolyte solution administered intracecally at two d...
Palomino Lago E, Roberts SJ, Orriss IR, Lumsden ES, Guest DJ.Bone is a dynamic tissue that is maintained through the co-ordination of bone resorption and bone formation. An imbalance of these processes can lead to bone disease. In vitro studies of osteoblasts can help to understand bone formation, but primary cells have a limited lifespan in culture. Herein, we report the successful generation of equine immortalized osteoblasts through the stable overexpression of human telomerase reverse transcriptase (hTERT) and Simian virus 40 (SV40) large T-antigen in osteoblasts isolated from trabecular bone taken from the third metacarpal of a two-year-old Thoroug...
Cortez JV, Cervi D, Ruiz AJ, Grupen CG.Nicotinic acid (NA) treatment during in vitro maturation (IVM) has been shown to elevate nicotinamide adenine dinucleotide (NAD) levels and improve oocyte developmental competence. Suboptimal equine oocyte IVM systems currently limit the efficiency of viable embryo in vitro production. This study evaluated NA supplementation during IVM for cloned equine embryo production, using oocytes from abattoir-sourced ovaries and live mares via ovum pick-up (OPU). Abattoir-derived oocytes (n = 694) were treated without or with 50 or 200 μM NA during the 18 h holding period (Pre-IVM). Next, OPU-deriv...
Meistro F, Rinnovati R, Blanc E, Berni P, Napoli S, Marcucci E, D'Angelo P, Ruggeri M, Spadari A, Gialletti R.Accurate early prognostic assessment is a central aspect of clinical decision-making in horses presenting with colic. Despite the availability of multiple clinical and laboratory parameters, reliable biomarkers that provide useful information at the time of admission remain limited. The erythrocyte sedimentation rate is a non-specific indicator of systemic inflammation that has recently been validated for automated point-of-care use in horses. Its behaviour in equine colic has not been previously characterised. This study included 85 horses admitted for colic at two university referral hospita...
Díaz-Alvarado T, Alcala-Canto Y, Elghandour MMMY, Salem MZM, Mariezcurrena-Berasain MD, Salem AZM.Gastrointestinal endoparasitic diseases, particularly those caused by some strongyles, represent one of the main health challenges in equine production. The prolonged use of synthetic anthelmintics has led to the development of resistance, which has prompted the search for more sustainable therapeutic alternatives. However, plant extracts rich in secondary metabolites are emerging as a promising option. Objective: The objective of this study was to evaluate the in vitro anthelmintic activity of aqueous extracts and bioactive compounds (total phenols and saponins) from Azadirachta indica and Cn...
Granello ME, Young JM, Cleff DB, Banks EBM, Trumble TN.To describe characteristics of postoperative peritoneal fluid following exploratory laparotomy for naturally-occurring gastrointestinal lesions in horses. Methods: Prospective, observational cohort study. Methods: A total of 26 client-owned horses that underwent exploratory laparotomy for naturally-occurring gastrointestinal lesions. Methods: Abdominocentesis was performed pre- or intraoperatively, and at three time points postoperatively (24, 72, and 168 h). Peritoneal lactate, total protein (TP), total nucleated cell count (TNCC), cytology, and systemic lactate were performed at each time ...
Tsopp E, Kilk K, Gambini A, Tvrdà E, Viljaste-Seera A, Tuppits U, Jaakma Ü.Equine in vitro embryo production (IVP) via ovum pick-up (OPU) and intracytoplasmic sperm injection (ICSI) has gained prominence in sport-horse breeding due to high success rates, efficient semen utilization, and year-round embryo production, including from mares with fertility challenges or postmortem. However, assessing viability of early-stage ICSI-produced equine embryos remains challenging due to subjective morphological evaluations and limited visibility of developmental features. Non-invasive metabolomic analysis of culture media offers a promising alternative to enhance embryo selectio...
Bindi F, de Marchi L, Elias-Cortajarena A, Sala G, Vitale V, Spadari A, Rinnovati R, Bonelli F, Sgorbini M.Colic is a major cause of morbidity and mortality in horses, with oxidative stress implicated in its pathophysiology. Objective: Evaluate biomarkers (BIOs) of oxidative stress and antioxidant defense in healthy horses and those with non-strangulating colic (NSC) and strangulating colic (SC) and assess correlations with survival. Methods: Seventy-one adult horses: 10 healthy and 61 colic-affected (42 NSC, 19 SC) admitted to 3 veterinary teaching hospitals. Methods: Prospective, multicenter cohort study. Blood samples were collected at admission (T0) and up to 96 h post-admission. Biomarkers me...
The ability of spermatozoa to withstand cryopreservation differs between stallions. While the underlying mechanisms of these differences are not fully understood, seminal plasma (SP) may play a crucial role in modulating sperm cryotolerance. Seminal plasma contains a heterogeneous population of extracellular vesicles (sEVs), nanometer-sized membrane particles that can transfer biomolecules to sperm modulating their function. This study aimed to isolate and characterize two sized-sEV subsets-small (S-) and large (L-)-from stallion SP, and to evaluate their involvement in sperm freezability. Sem...
Enkhtuul B, Khurtsbaatar O, Lkham B, Agiimaa T, Unenbat J, Buyanbadrakh B, Ninjbulgan S, Liushiqi B, Suzuki Y, Kimura T, Batbaatar V.Glanders, a zoonotic disease caused by , has re-emerged in Mongolia after decades of control. We conducted nationwide serosurveillance from 2021 to 2024 to assess the prevalence and geographic distribution of glanders. Using random 3-stage cluster sampling, we collected 3,001 horse serum samples from 332 herds. Those herds were in 46 soums (administrative districts equivalent to counties) in 13 provinces and in 3 districts in Ulaanbaatar. Sera were tested using the complement fixation test, and positive results were confirmed by immunoblotting and ELISA. Overall seroprevalence was 1.03% (95% C...
Normélia P Morais A, Gastal GDA, Aguiar FLN, Hyde KA, Gastal MO, Lima LF, Ñaupas LVS, Flávia B Silva A, Becerra VAB, Gheno L, Curcio BR....Understanding how to effectively isolate a large number of healthy follicles is essential for increasing success rates in ARTs, such as in vitro culture and cryopreservation. Late secondary (preantral) and early tertiary (antral) follicles isolated from the intermediary portion of equine ovarian tissue sliced at two different thicknesses (0.25 mm vs. 0.50 mm) were characterized according to regional distribution (dorsal, medial, and ventral), morphofunctional features, and epigenetic profiles. After isolation via microdissection, follicles were evaluated for membrane integrity (trypan blue), v...
Fernández-Villa C, Miranda M, Rigueira L, Martinez L, Villanueva B, Freire S, López-Alonso M.Serum mineral concentrations are key indicators of nutritional status, metabolic function and health in horses; however, contemporary reference intervals (RI) derived from large, well-characterised populations remain scarce. Objective: To establish serum RI for macro- and microminerals in clinically healthy horses in Northwestern Spain and to evaluate the influence of sex, age, breed and diet on mineral status. Methods: Blood samples were collected from clinically healthy horses (n = 211), and serum concentrations of essential macrominerals (calcium [Ca]; phosphorous [P]; magnesium [Mg]; sodiu...
Nunes MM, Gurgel JRC, Munhoz ML, de Giuli V, Carneiro GF, Miragaya MH, Gallelli MF, Guimaraes JD.The aim of this study was to assess the effect of lyophilized freezing extenders, which can be stored at room temperature, on stallion post-thaw sperm total motility (TM). Ejaculates of 28 stallions were frozen with four different extenders: two commercial freezing extenders offered worldwide and two novel lyophilized extenders (STAR and MX3), and two different cryopreservation protocols (CP1 with an equilibration period of 20 min. and CP2 with an equilibration period of 60 min.). The TM was assessed after thaw. Mean TM did not show significant differences between cryopreservation protocols wi...
Kushida C, Tamura N, Kasashima Y, Sato K, Arai K.In equine regenerative medicine using bone marrow-derived mesenchymal stem/stromal cells (BM-MSC), the importance of the quality management of BM-MSC has been widely recognized. However, there is little information concerning the relationship between cellular senescence and the stemness in equine BM-MSC. In this study, we showed that stemness markers (NANOG, OCT4, SOX2 and telomerase reverse transcriptase) and colony forming unit-fibroblast apparently decreased accompanied with incidence of senescence-associated β-galactosidase-positive cells by repeated passage. Additionally, we suggested th...
Gray B, Lubbock K, Love C, Ryder E, Hudson S, Scarth J.The analytical approaches taken by laboratories to implement robust and efficient regulation of horseracing medication and doping control are complex and constantly evolving. Each laboratory's approach will be dictated by differences in regulatory, economic and scientific drivers specific to their local environment. However, in general, laboratories will all be undertaking developments and improvements to their screening strategies in order to meet new and emerging threats as well as provide improved service to their customers. In this paper, the published analytical advances in horseracing me...
Ramírez-Agámez L, Castaneda C, Hernández-Avilés C, Grahn RA, Raudsepp T, Love CC.Two methods for preimplantation genetic testing (PGT) have been described for equine embryos: trophoblast cell biopsy (TCB) or blastocoele fluid aspiration (BFA). While TCB is widely applied for both in vivo- and in vitro-produced embryos, BFA has been mostly utilized for in vivo-produced embryos. Alternative methods for PGT, including analysis of cell-free DNA (CFD) in the medium where in vitro-produced embryos are cultured, have been reported in humans but not for equine embryos. In Experiment 1, in vivo- (n = 10) and in vitro-produced (n = 13) equine embryos were subjected to BFA, cultu...
Caldevilla M, Ferrante A, Neild DM.The addition of antioxidants to cryopreservation media reportedly improves sperm post-thaw quality and reproductive performance after artificial insemination. Therefore, the objectives of this study were to evaluate if the addition of L-carnitine and pyruvate to freezing media, or their addition to samples after thawing, improves the post-thaw quality of equine spermatozoa. Thus, in Experiment 1, stallion semen samples were cryopreserved in: (1) EDTA-glucose-based extender with 20% egg yolk and 5% dimethylformamide (EDTA control); (2) skim milk-based extender with 20% egg yolk and 5% dimethylf...
Medica AJ, Gibb Z, Aitken RJ.In vitro semen purification techniques have been developed that seek to mimic the in vivo selection process in order to generate the highest possible chance of oocyte fertilization following artificial insemination. Numerous methods have been developed to isolate functional spermatozoa for artificial insemination, yet only one method, single-layer centrifugation using commercial preparations like EquiPure, has been widely employed. In this study, we have introduced a novel approach for isolating spermatozoa and compared their quality to those isolated using EquiPure. The AI port system (Mempha...
Dias Moreira AS, Weng HY, Hostnik LD, Beasley EM, Peek SF, Munsterman AS.To compare glucose measurements from capillary and venous blood samples using a point-of-care (POC) glucometer with a standard laboratory (colorimetric, glucose oxidase) assay (LABGLU) in a population of hospitalized, neonatal foals. Methods: Multicenter, prospective, experimental study, conducted between March 2019 and June 2020. Methods: Four university teaching hospitals and 1 private referral hospital. Methods: Fifty-four hospitalized neonatal (≤30 days of age) foals. Methods: Simultaneous capillary (muzzle, POCMUZ) and venous (jugular, POCJUG) blood samples were obtained to determine P...
Hoerdemann M, Sahoo DK, Allbaugh RA, Kubai MA.To assess if an affordable, commercially available 275 nm ultraviolet C (UV-C) device can inhibit fungal pathogens associated with equine keratomycosis and to establish the optimal exposure time/dose required in vitro. Methods: Fungal inhibition zone surface areas of isolates from two fungal genera (Aspergillus and Fusarium spp.) were compared in triplicates after UV-C exposure at distances of 10, 15, and 20 mm with single or repeat doses (4 h apart) of 5, 10, 15, or 30 s duration after 24, 48, and 72 h of incubation. Inhibition zones were microscopically assessed, and regrowth chec...
Spacek SG, Hernández-Avilés C, Ramírez-Agámez L, Pearson M, Love CC.Cryopreservation of stallion epididymal sperm has become a common clinical procedure after routine castration, euthanasia, or acute death. Unique features of epididymal sperm compared to ejaculated sperm include the requirement to remove sperm from the cauda epididymis, lack of exposure to seminal plasma, and potential recovery of large sperm numbers. In this study, the effect of the flushing technique (Extender (INRA) vs. AIR), freezing extender (LE, CMLE, MFR5, CFR5, or BOTU), and the concentration at which sperm are cryopreserved (200, 400, or 800 × 10 sperm/mL) on post-thaw epididymal s...
Vieira GS, Peneiras ABV, Soares TSC, Alonso MA, Curcio BR, Fernandes CB, Oliveira RA.Advancements in equine neonatology enable continuous monitoring of mares throughout pregnancy. However, reference values for oxygenation and biochemical parameters in newborn foals remain limited. Objective: This study aimed to assess the oxygenation status of healthy Mangalarga Marchador foals from birth to 24 h postpartum by evaluating blood gas analysis, Apgar scores, and biochemical and hematological parameters. Methods: Thirty healthy Mangalarga Marchador foals were evaluated. Blood samples were collected from the umbilical artery at birth (T0) and from the dorsal metatarsal artery at 15 ...
