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Topic:Analytical Methods
Analytical methods in equine research encompass a variety of scientific techniques and tools used to study and evaluate different aspects of horse health, performance, and physiology. These methods help advance our understanding of equine biology, diagnosing conditions, and improving management practices. Common analytical methods include molecular techniques like PCR and ELISA for detecting pathogens and measuring biomarkers, imaging technologies such as ultrasound and MRI for assessing musculoskeletal health, and statistical models for analyzing genetic data and performance metrics. This page compiles peer-reviewed research studies and scholarly articles that explore the development, application, and impact of various analytical methods in equine science.
Characterization of Capillary Zone Electrophoresis in Horses and Donkeys. There are no capillary zone electrophoresis (CZE) comparative studies between horses and donkeys. Our objectives were to compare CZE results between both species, establish reference intervals (Ris), study age- and sex-related changes, determine the prevalence of bisalbuminemia, and correlate spectrophotometric albumin results with those calculated based on two different CZE methods. We performed CZE in 50 healthy horses and 49 healthy donkeys. There were significant differences ( < 0.03) in every protein fraction percentage and concentration (except for the β-globulins percentage) between...
Investigation of Metformin Contamination in Horses: Implications for Environmental Exposure and Doping Control. Metformin is a diabetes medication with minimal therapeutic efficacy in veterinary medicine. However, recently there have been several adverse analytical findings in regulatory samples collected from racehorses. Metformin is commonly prescribed to humans, is excreted unchanged in urine, and is resistant to environmental degradation, raising concerns with respect to environmental contamination. In this study, six horses were exposed to shavings contaminated with metformin-positive urine at concentrations of 10, 135, and 270 μg/mL. Blood (plasma, serum, whole blood, and red blood cells) and u...
Synchronization dynamics between rider and saddle motion across gaits revealed by IMU analysis. Effective rider-horse coordination is essential for performance and welfare, yet objective, field-based quantification of rider-saddle synchronization across gaits remains limited. Objective: To quantify dynamic synchronization between rider motion and saddle motion (as a proxy for horse trunk oscillations) across walk, trot, and canter using a minimal inertial measurement unit (IMU) setup, and to determine whether synchronization differs between rider body segments. Methods: Two experienced riders (8-20 years riding experience) rode their own or familiar warmblood horses (n = 2; age 6-14 year...
Pharmacokinetics of Diclazuril in the Plasma and Cerebrospinal Fluid of Healthy Adult Horses After a Single Oral Dose Administered as a Pelleted Topdressing. Equine protozoal myeloencephalitis can cause acute infections with rapid onset of neurologic signs, necessitating immediate empiric therapy with antiprotozoal medication. The objective of the study was to identify when concentrations of diclazuril reached the MIC of Sarcocystis neurona in CSF (1 ng/mL) of healthy adult horses after a single oral dose. Six healthy adult horses were used. Blood and CSF were collected from indwelling intravenous jugular catheters and intrathecal catheters in the lumbosacral space prior to drug administration and then at 1-, 4-, 8-, 12-, 16-, 24-, 48-, 72-, 96-,...
LC-HRMS detection of three ionizable lipids in equine plasma: surrogate markers for long-term LNP-mediated gene-doping surveillance. Lipid nanoparticles (LNPs) are essential delivery vehicles for messenger ribonucleic acid (mRNA)-based therapies. However, their potential misuse as gene-doping agents in horse racing and equestrian sports necessitates robust analytical surveillance. This study presents a highly sensitive, Orbitrap technology-driven liquid chromatography-high-resolution mass spectrometry (LC-HRMS) method for the simultaneous detection of three clinically relevant ionizable lipids: ALC-0315, SM-102, and DLin-MC3-DMA (MC3), which are critical components of approved RNA therapeutics and vaccines. Following a modi...
Lacrimal caruncula thermography overestimates rectal temperature in horses: Influence of camera type and measurement distance. Rectal temperature (RT) is the gold standard for assessing body temperature in horses, but handling and welfare concerns limit its use. Thermography of the lacrimal caruncula (LC) has been proposed as a noncontact alternative, although its accuracy may be influenced by device characteristics and measurement distance. Objective: To evaluate agreement between LC thermography and RT in horses, compare two infrared thermal cameras and determine the effect of camera-to-subject distance on measurement bias and variability. Methods: LC temperature was recorded at 0.5, 1 and 2 using two cameras (IR Fl...
Sensitive and selective quantification of elcatonin in equine plasma and urine using LC-FAIMS-MS/MS. The use of drugs for enhancing bone strength is prohibited under international regulations governing horse racing and equestrian sports. Elcatonin, a synthetic peptide with analgesic and bone-strengthening properties, is routinely monitored for compliance. However, its cyclic structure and high molecular weight hinder reliable detection. In this study, we developed and validated a sensitive liquid chromatography-high-field asymmetric waveform ion mobility spectrometry-tandem mass spectrometry method for quantifying elcatonin in equine plasma and urine. Sample preparation was optimised via weak...
Telomere-to-telomere Genome Assembly of Equus kiang (Tibetan wild donkey). The Tibetan wild donkey (Equus kiang) is an endemic equid species inhabiting the Qinghai-Tibet Plateau, exhibiting remarkable chromosomal diversity within the Equus genus. Here we present a chromosome-level, telomere-to-telomere genome assembly for E. kiang generated using Illumina short reads, PacBio HiFi long reads, and Hi-C sequencing data. The final assembly spans 4.02 Gb and comprises 27 pseudochromosomes. Specifically, 8 chromosomes are gapless, 8 chromosomes have one gap, 7 chromosomes have 2-4 gaps, and the remaining 4 chromosomes have five or more gaps. Benchmarking Universal Single...
Comparison of in vitro degradation of immunoreactive adrenocorticotropic hormone and insulin in equine blood measured by two immunoassay methods. This study compared the in vitro stability of equine plasma adrenocorticotrophic hormone (ACTH) and serum insulin measured by chemiluminescent assay (CLA; Siemens Immulite 2000XPi) and immunofluorescent assay (IFA; Tosoh AIA-900) after storage at 4°C or room temperature (approximately 21°C) for clinically relevant periods. Fresh equine EDTA blood for ACTH and coagulated blood for insulin were aliquoted and analysed immediately (T0) and after storage. ACTH was assessed at 24, 48 and 72 hours; insulin was assessed at 24 hours, 72 hours and 7 days. Percentage recovery relative to T0 was analyse...
Clodronate disodium reduced TRAP5b independent of affecting other systemic biomarkers of bone resorption or formation in yearling, exercising Quarter Horses. Clodronate disodium (CD), a bisphosphonate, modulates bone metabolism. Though extra-label use in juvenile horses is anecdotally reported, impacts on skeletal development are unknown. The objective was to determine the effects of CD on systemic markers of bone turnover in yearling horses undergoing exercise, hypothesizing that biomarkers of bone resorption would decrease while biomarkers of bone formation would not change, and that repeat CD treatments would have greater effect. To test this, 32 yearling Quarter Horses were used in a 168-d trial. Horses were stratified by age (500 ± 13 d),...
Validation of a stall-side immunoglobulin assay for use in equine reproductive management. Equine foals receive IgG from mare colostrum through passive transfer. Failure of passive transfer (FPT) is a significant risk to the foal's life, leaving them vulnerable to infection and sepsis. Radial Immunodiffusion (RID) and immunoturbidimetric assays quantify IgG present in a foal sample but require a laboratory to complete. Accurate, reproducible, stall-side testing to rapidly quantify IgG would allow for expedited clinical decisions, with potential to improve equine foal care and survival. Objective: To evaluate the analytical and clinical performance of a stall-side IgG lateral-flow te...
Comparison Between Inertial Sensor and Video-Based Detection of Spatiotemporal Limb Movement Parameters During Equine Swimming. Equine swimming is increasingly used for injury prevention and rehabilitation, but objective analysis of movement during swimming remains limited compared to land-based locomotion. Spatiotemporal parameters are essential for evaluating therapeutic outcomes, yet capturing these parameters is technically challenging due to difficulties in observing limb motion in water. Inertial sensors, already widely applied in equine science, offer a promising solution for measuring swimming kinematics objectively. The objective of this study was to evaluate the reliability of inertial sensors placed on equin...
A scoping review of neuroendocrinological biomarkers with potential for use in equine behavioral medicine practice. BACKGROUND: Equine emotion and welfare are of increasing concern to the general public, equestrians and researchers. There is growing demand for more objective indicators of specific emotional states, as well as the well-being of horses in a variety of contexts. Such markers may also assist in both the diagnosis and selection of biological therapies within equine behavioral medicine. RESULTS: This scoping review summarizes and appraises the published findings relating to relevant biological measurements from the last two decades. Particular attention is given to methodological, subject-related...
Ertugliflozin decreases the insulin spike in non-insulin dysregulated standardbred horses following intra-articular triamcinolone administration. Horses with equine metabolic syndrome (EMS) are at an increased risk of laminitis following corticosteroid use. Ertugliflozin reduces hyperinsulinemia in horses with EMS, and therefore may be a management option to reduce the insulin spike seen post corticosteroid use. It was hypothesised that oral ertugliflozin, administered before and after intra-articular (IA) triamcinolone acetonide (TA), will significantly lower the insulin response in comparison to placebo. A randomised, blinded, placebo-controlled, crossover design was performed on eight adult standardbred geldings with no history of la...
Application of 3D scanning for modeling and morphometric assessment of Mangalarga Marchador Horses. Morphometric traits are essential for assessing conformation, locomotion, and selection in equine breeding programs. However, manual measurements are laborious, require physical contact, and may cause stress to animals, highlighting the need for alternative technologies. Objective: This study evaluated the use of a portable 3D scanner for three-dimensional modeling and morphometric assessment of Mangalarga Marchador horses. Methods: Fifty-two adult horses were measured using manual and 3D scanning methods to assess ten morphometric traits: withers height (WH), hip height (HH), head length (HL)...
Transference of diagnostic thresholds for equine plasma ACTH from the Immulite 1000 to Immulite 2000XPi and Tosoh AIA systems. Diagnosis of PPID in horses relies on measurement of plasma adrenocorticotrophic hormone (ACTH) concentrations. Because measured ACTH values vary between analytical platforms, and published diagnostic thresholds apply almost exclusively to the Immulite 1000 chemiluminescent assay (CLA), there is a need for validated, assay-specific thresholds for contemporary analysers. This study aimed to transfer and validate established Immulite 1000 diagnostic thresholds for plasma ACTH to three contemporary systems (Immulite 2000XPi, Tosoh AIA-900, and Tosoh AIA-360) using the Clinical Laboratory Standard...
Integrating clinical and rna-seq findings in a mare with recurrent endocrinopathic laminitis linked to equine metabolic syndrome. Equine metabolic syndrome (EMS) is characterized by insulin dysregulation and obesity, which increase the risk of the development of hyperinsulinemia-induced laminitis (HAL). The present report describes a case of severe endocrinopathic laminitis associated with EMS, refractory to treatment. A 5-year-old mare with recurrent HAL presented transient relief upon a low nonstructural carbohydrate diet and conventional therapy. An SGLT2 inhibitor was introduced to reduce resting insulin levels, and improvement was observed until October 2024, when the most severe episode started. Despite adding perg...
Erythrocyte sequestration of metformin in horses: impact on matrix-specific pharmacokinetics and detection windows. BACKGROUND: Metformin is used for the treatment of type 2 diabetes and is one of the most prescribed medications in human medicine. It is less commonly prescribed in equine medicine, and its use is tightly regulated in several performance horse disciplines. In horseracing, it is considered a banned substance. A previous pharmacokinetic study of metformin in horses demonstrated a prolonged, unpredictable elimination phase. In the current study it was hypothesized that this was due to sequestration of metformin in a “deep” compartment, specifically red blood cells. This could result in pharm...
Genetic diversity in the Criollo Argentino horse from SNP array data. The Criollo Argentino horse is a national symbol and a heritage breed that represents both cultural identity and a key component of traditional production systems in Argentina. This study aimed to assess the genetic differentiation of the Criollo Argentino relative to other horse breeds and to infer potential factors shaping its diversity through the genomic analysis of 24 representative animals. Samples were genotyped using a medium-density SNP array containing 65,157 markers, and genetic diversity analyses were conducted across autosomal SNP sets filtered under different criteria. Contempora...
Evaluating Protein Enrichment Methods to Improve Biomarker Discovery in Equine Cerebrospinal Fluid. Cerebrospinal fluid (CSF) is a valuable source of biomarkers for neurological diseases, but detection of low-abundance proteins is often masked by highly abundant proteins. Enrichment strategies can enhance proteomic coverage and improve biomarker discovery, yet comparative evaluations of such methods in equine CSF are limited. Objective: This study compared the ProteoMiner Small-Capacity Enrichment Kit and the PreOmics Enrich-iST Kit for their ability to deplete high-abundance proteins and enhance detection of low-abundance proteins relevant to neuropathology. Methods: Equine CSF samples were...
Diagnostic Threshold Identification for Equine Laminitis Using Smoothed Receiver Operating Characteristic Analysis. Radiographic measurement parameters play a key role in diagnosing acute and subacute laminitis in horses, with diagnostic thresholds typically derived from empirical receiver operating characteristic (ROC) analysis. However, these methods often produce unstable results, particularly in small or noisy datasets. This study applies nonuniform rational B-spline (NURBS)-based ROC smoothing to radiographic data from laminitic and healthy horses to evaluate its effectiveness in refining diagnostic thresholds. Thresholds are determined using Youden's index based on NURBS-smoothed ROC curves and compar...
Objective gait analysis methods for assessing movement asymmetry in the equine patient. To demonstrate the uses, data interpretation, and limitations of objective gait analysis methods in equine practice. Unassigned: Teaching and client-owned horses. Unassigned: Informed owner consent was obtained for all animals used in the making of this technical tutorial video. Primary literature relevant to this video was reviewed from January 2016 to January 2026. Images and data were collected from teaching horses and client-owned horses. Unassigned: Objective gait analysis provides accurate and repeatable data on the locomotion of a horse; however, inaccuracies may occur, especially in ca...
Prevotella mikamonis sp. nov., isolated from equine clinical specimens. In this study, five strains of anaerobic, Gram-negative, rod-shaped bacteria were characterized phenotypically, biochemically and genotypically. These strains were clinically isolated from horse specimens in Japan. Phylogenetic trees constructed based on 16S rRNA gene sequence comparisons showed distinct clustering of the five strains with the type strains of closely related species. The genomic DNA G+C content was 46.7 mol%. The major cellular fatty acids identified were C, 3-OH-C and 3-OH-iso-C. Whole-genome comparisons based on average nucleotide identity using blast (ANIb) and digital DNA...
Analytical Challenges and Emerging Detection Strategies of Synthetic Cannabinoids in Horse Doping Control. Synthetic cannabinoids (SCs) were originally synthesized to advance the understanding of the endocannabinoid system, facilitate disease research, and support the development of novel therapeutic agents. Compared with tetrahydrocannabinol, these compounds exhibit substantially higher psychoactive potency and enhanced receptor-binding affinity. The rapid and continuous evolution of SC derivatives presents significant challenges for analytical laboratories and increases the risk of misuse. Furthermore, SCs may be exploited to alter performance in both human and animal sports. The scarcity of comp...
The transition from winter to spring has an impact on the airway metabolome profile of asthmatic horses. Equine asthma is a chronic, non-infectious inflammatory disease of the lower airways in horses, classified as mild to moderate (MEA) or severe (SEA). Its pathogenesis is not fully understood and is influenced by environmental and seasonal factors. In this cross-sectional study, seasonal effects on the bronchoalveolar lavage fluid (BALF) metabolome were investigated in asthmatic and non-asthmatic horses. The metabolome of 230 BALF samples from horses across different seasons, classified as cytologically unremarkable (CUA), MEA, or SEA, was analyzed using proton nuclear magnetic resonance spectr...
Saliva and plasma metabolome analysis during the five days before foaling in the mare. Saliva is a relevant biofluid for real-time welfare-friendly monitoring of systemic events in animals, because some bioanalytes have a systemic origin and its collection is painless, stress-free and non-invasive. Our aim was to analyze the metabolome of equine saliva during the five days before foaling, with a focus on identifying metabolites whose level significantly changed before parturition, that could be potential salivary biomarkers of the onset of parturition. We compared the saliva and plasma metabolomes to investigate their relationship. Saliva and blood samples were collected from tw...
Pharmacokinetics of Intramuscularly Administered Ketamine at Subanaesthetic Dosage in Horses. Subanaesthetic doses of ketamine (0.5 mg kg) provide analgesia in several species, but there is limited information on the pharmacokinetics and clinical effects of intramuscularly administered ketamine in horses. This study investigated the pharmacokinetics of ketamine and norketamine, and evaluated clinical effects, ataxia, and vital parameters, following intramuscular and intravenous administration of 0.5 mg kg ketamine in nine healthy horses, using a randomized two-period crossover design with a 1-week washout period. Plasma concentrations were analyzed using Ultra-High Performance ...
Sex determination and genetic screening of equine embryos with whole genome amplification and real-time PCR. Reliable pre-implantation sex determination and genetic screening enables informed embryo transfer decisions in equine breeding while avoiding later interventions. We developed a streamlined workflow that couples rapid whole genome amplification (WGA) with a multiplex real-time PCR targeting ETSTY5 as a Y-specific marker and UBC as an autosomal control. On purified equine DNA, sex was correctly assigned down to 10 pg gDNA and to a single fibroblast cell. Direct testing of embryo biopsies without WGA yielded inconsistent results, whereas introducing a short WGA step produced tight allelic-discr...
