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Topic:Immunofluorescence Assay
Immunofluorescence assay (IFA) is a laboratory technique used to detect and visualize specific antigens or antibodies in equine tissue samples or bodily fluids. This method employs fluorescent-labeled antibodies to bind target molecules, allowing for the observation of fluorescence under a microscope. In horses, IFA is employed in various research and diagnostic applications, including the study of infectious diseases, immune responses, and cellular localization of proteins. The technique provides valuable insights into the distribution and expression of specific proteins within equine cells and tissues. This page aggregates peer-reviewed research studies and scholarly articles that explore the methodology, applications, and advancements of immunofluorescence assay in equine research.
A cross-sectional study of Leishmania spp. in draft horses from the Distrito Federal, Brazil: Seroprevalence, spatial distribution, and associated factors. Equine leishmaniasis is a neglected tropical disease caused by the protozoan of the Leishmania genus, and it has been reported in several countries around the world, especially Brazil. Therefore, the present investigation aims to conduct a cross-sectional study to determine the prevalence, spatial distribution, and associated factors with seropositivity for Leishmania spp. in draft horses from the Distrito Federal, Brazil. The serological survey was conducted on 411 animals, employing the Indirect Immunofluorescence Test (IFA) and Enzyme-Linked Immunosorbent Assay (ELISA). The Kappa (κ) and g...
RNA sequencing and immunofluorescence of the myotendinous junction of mature horses and humans. The myotendinous junction (MTJ) is a specialized interface for transmitting high forces between the muscle and tendon and yet the MTJ is a common site of strain injury with a high recurrence rate. The aim of this study was to identify previously unknown MTJ components in mature animals and humans. Samples were obtained from the superficial digital flexor (SDF) muscle-tendon interface of 20 horses, and the tissue was separated through a sequential cryosectioning approach into muscle, MTJ (muscle tissue enriched in myofiber tips attached to the tendon), and tendon fractions. RT-PCR was performed...
Theileria equi and Babesia caballi in horses in the Czech Republic. Theileria equi and Babesia caballi are protozoan agents causing equine piroplasmosis, endemic in countries all over the world. The aim of this study was to detect antibodies to T. equi and B. caballi in horses in the Czech Republic and to investigate the origin of the infection. Blood sera from 711 horses were examined with competitive enzyme-linked immunosorbent assay; positive samples were verified with indirect fluorescence immunoassay. Antibodies to T. equi and B. caballi were detected in eight (1.1%) and three (0.4%) horses, respectively. Infection with T. equi was confirmed by PCR and se...
Serosurvey for equine piroplasms in horses and donkeys from North-Western Nigeria using IFAT and ELISA. Equine piroplasmosis is caused by apicomplexan parasites, namely, and , which are transmitted to equids principally through ticks. To ascertain the exposure of equines to agents of equine piroplasms, we tested serum samples collected from horses (n = 272) and donkeys (n = 170) in North-Western Nigeria for the presence of antibodies against and using IFAT and ELISA. The seroprevalence of in the horses determined using IFAT and ELISA was 48.89% and 45.96%, respectively, while for , it was 6.3% and 0.4%, respectively. For , the seroprevalence based on IFAT and ELISA results in donkeys was...
Seroepidemiology of Sarcocystis neurona and Neospora spp. in horses, donkeys, and mules from Colombia. Sarcocystis neurona and Neospora spp. are related protozoa that can cause equine protozoal myeloencephalitis (EPM). The present study aimed to determine the frequency of antibodies to these parasites in 649 equids (351 horses, 267 donkeys, and 31 mules) from six departments in the North and Northwest of Colombia. For this purpose, the indirect fluorescent antibody test (IFAT) was used for detecting antibodies against S. neurona and Neospora spp. with a cut-off point of 1:20 and 1:50, respectively. A binomial logistic regression model was selected to predict variables associated with exposure. ...
Development of a constant pressure perfused ex vivo model of the equine larynx. Distal axonopathy is seen in a broad range of species including equine patients. In horses, this degenerative disorder of the recurrent laryngeal nerve is described as recurrent laryngeal neuropathy (RLN). The dysfunctional innervation of the cricoarytenoideus dorsalis muscle (CAD) leads to a loss of performance in affected horses. In general, ex vivo models of the larynx are rare and for equine patients, just one short report is available. To allow for testing new therapy approaches in an isolated organ model, we examined equine larynges in a constant pressure perfused setup. In order to chec...
ANCHOR-tagged equine herpesvirus 1: A new tool for monitoring viral infection and discovering new antiviral compounds. Equine herpesvirus 1 (EHV-1) is a causative agent of respiratory disorders, abortion and myeloencephalopathy in horses and has an important impact on equine health and economy. Several bacterial artificial chromosomes have already been developed and enabled identification and functional characterization of EHV-1 genes. Unfortunately, little is known about its replication. Here, the ANCHOR system was inserted by targeted homologous recombination into the equine herpesvirus genome. This insertion led to the conversion of EHV-1 DNA to auto-fluorescent spots easily detectable by fluorescence micro...
Serological and Molecular Prevalence of Babesia caballi in Apparently Healthy Horses in Israel. Babesia caballi is a tick-borne hemoparasite of equines and one of the causative agents of equine piroplasmosis, which poses a great concern for the equine industry regarding animal welfare and international horse movement. The parasite is endemic in Israel; however, its seroprevalence in the area was never evaluated due to antigenic heterogenicity in the gene used in the commercially available kit. Blood samples were collected from 257 horses at 19 farms throughout the country and screened for the presence of anti-B. caballi antibodies via an indirect immunofluorescent antibody test (IFAT) an...
The first serological evidence of Anaplasma phagocytophilum in horses in Slovakia. Anaplasma phagocytophilum is the causative agent of granulocytic anaplasmosis. It affects humans and several wild and domesticated mammals, including horses. The aim of our study was a preliminary survey of the occurrence of these re-emerging pathogens in horses in Slovakia. The sera from 200 animals of different ages and both sexes were tested for the presence of A. phagocytophilum antibodies by indirect immunofluorescence assay. Subsequently, detection of the 16S rRNA gene fragment of A. phagocytophilum was attempted by polymerase chain reaction (PCR) in each blood sample. Our results confir...
Serological evidence of Rickettsia in horses from a semi-arid Brazilian region. Brazilian spotted fever (BSF) is a common tick-borne disease caused by Rickettsia rickettsii. Horses are the primary hosts of the main vector, Amblyomma sculptum, and are considered efficient sentinels for circulation of Rickettsia. Therefore, the aim of this study was to detect antibodies reactive to R. rickettsii antigens in horses from a non-endemic area in the north-central region of Bahia state, Brazil. Blood samples and ticks were collected from 70 horses from the municipalities of Senhor do Bonfim, Antônio Gonçalves, Pindobaçu, and Campo Formoso. The sera obtained were tested by an i...
Detection of Ehrlichia sp. in Amblyomma sculptum parasitizing horses from Brazilian Pantanal wetland. Bacteria of the genus Ehrlichia are transmitted by ticks and also are an important cause of infection in wild and domestic mammals. Infection with Ehrlichia spp. has been reported in horses, especially in the USA, Nicaragua and Brazil. In this study, we report the parasitism by Amblyomma sculptum, Rhipicephalus microplus and Dermacentor nitens ticks in horses from a ranch located in south Pantanal wetland. Molecular and serological analyzes to determine infection by Ehrlichia spp. in horses and their respective ticks were carried out. A total of 12 horses were submitted to blood collection to ...
Expression of Proteinase-Activated Receptor 2 During Colon Volvulus in the Horse. Large colon volvulus in horses is associated with a poor prognosis, especially when ischemic-reperfusion injury of the affected intestinal tract develops. Proteinase-activated receptor 2 (PAR2) plays an important role in the pathogenesis of inflammation in the gastrointestinal tract. The aim of this study was to evaluate the distribution and expression of PAR2 in colonic pelvic flexure of horses spontaneously affected by large colon volvulus (CVH group). Eight horses admitted for severe abdominal colon volvolus and which underwent surgery were included. Colon samples were collected after enter...
In Vitro Induction of Pluripotency from Equine Fibroblasts in 20% or 5% Oxygen. The cellular reprogramming into pluripotency is influenced by external and internal cellular factors, such as culture conditions (e.g., environmental oxygen concentration), and the aging process. Herein, we aimed to generate and maintain equine iPSCs (eiPSCs) derived from fibroblasts of a horse older than 20 years and to evaluate the effect of different levels of oxygen tension (atmospheric 20% O, 5% O, or 20% to 5% O) on these cells. Fibroblasts were reprogrammed, and putative eiPSCs were positive for positive alkaline phosphatase detection; they were positive for pluripotency-related genes ...
Rickettsial infection in equids, opossums and ticks in the municipality of Monte Mor, state of São Paulo, Brazil. The aim of this study was to investigate rickettsial infection in equids, opossums and ticks in the municipality of Monte Mor, a place where a Brazilian spotted fever case occurred in 2005. In addition, characteristics possibly associated with seropositivity in horses were analyzed. Serum samples from horses, mules and opossums (Didelphis albiventris) were subjected to indirect immunofluorescence assay (IFA) against Rickettsia rickettsii. The ticks collected from the animals were identified and Amblyomma sculptum ticks from the equids were tested using PCR for Rickettsia spp. Anti-R. rickettsi...
PD-L1 expression in equine malignant melanoma and functional effects of PD-L1 blockade. Programmed death-1 (PD-1) is an immunoinhibitory receptor expressed on lymphocytes. Interaction of PD-1 with its ligand PD-ligand 1 (PD-L1) delivers inhibitory signals and impairs proliferation, cytokine production, and cytotoxicity of T cells. In our previous studies, we have developed anti-bovine PD-L1 monoclonal antibodies (mAbs) and reported that the PD-1/PD-L1 pathway was closely associated with T-cell exhaustion and disease progression in bovine chronic infections and canine tumors. Furthermore, we found that blocking antibodies that target PD-1 and PD-L1 restore T-cell functions and cou...
Reactivity of Horse Sera to Antigens Derived From Sarcocystis falcatula-Like and Sarcocystis neurona. and are protozoan parasites endemic to the Americas. The former is the major cause of equine protozoal myeloencephalitis, and the latter is associated with pulmonary sarcocystosis in birds. The opossum is the definitive host of these parasites in North America. Four species are found in Brazil, and in most reports in this country, species shed by opossums have been classified as -like. It is unknown whether reports on -seropositive horses in Brazil are also derived from exposure of horses to -like. The aim of this study was to test the sera reactivity of 409 horses in Brazil using antigen...
Detecting antibodies to Leishmania infantum in horses from areas with different epizooticity levels of canine leishmaniosis and a retrospective revision of Italian data. Leishmania infantum is a vector-borne pathogen endemic in countries in the Mediterranean basin, including Italy. Dogs act as the primary reservoir for this parasite, but other animal species may also be infected. Low-to-moderate seroprevalence levels of infection have been reported in apparent healthy equine populations in southern Europe, reinforcing the importance of exploring those species, including horses, that act as a food source for vectors and may thus participate in the epizoological scenario of canine leishmaniosis (CanL) and zoonotic visceral leishmaniosis (ZVL). Since little is kn...
The expression of androstenone receptor (OR7D4) in vomeronasal organ and olfactory epithelium of horses. Androstenone is the first mammalian steroidal pheromone to be identified. Pheromones are chemicals that animals use to communicate within a species. Pheromone detections are related to vomeronasal organ (VNO) and olfactory epithelium (OE) in mammals. Olfactory Receptor Family 7 Subfamily D Member 4 (OR7D4) is an odorant receptor that responds to androstenone. Several studies indicated that spray with androstenone changes behaviors of the boar and dogs. However, the expression of OR7D4 in VNO and OE was not reported in mammals except human. Thus, the main objectives of this study were to invest...
Seroprevalence of Antibodies Against Anaplasma Phagocytophilum and Borrelia Burgdorferi in Horses (Equus Caballus) from Northern Algeria. Horses (Equus caballus) are susceptible to tick-borne diseases. Two of them, Lyme borreliosis due to Borrelia burgdorferi and granulocytic anaplasmosis due to Anaplasma phagocytophilum were investigated in Algerian horses. The diseases have been less extensively studied in horses and results pertinent to Algeria have not been published. Methods: Blood samples were obtained from 128 horses. IgG antibodies directed against Anaplasma phagocytophilum and Borrelia burgdorferi were detected by an indirect immunofluorescence antibody test (IFAT) and ELISA. The potential effects of age, gender, breed,...
Type I Collagen Suspension Induces Neocollagenesis and Myodifferentiation in Fibroblasts In Vitro. The ability of a collagen-based matrix to support cell proliferation, migration, and infiltration has been reported; however, the direct effect of an aqueous collagen suspension on cell cultures has not been studied yet. In this work, the effects of a high-concentration aqueous suspension of a micronized type I equine collagen (EC-I) have been evaluated on a normal mouse fibroblast cell line. Immunofluorescence analysis showed the ability of EC-I to induce a significant increase of type I and III collagen levels, parallel with overexpression of crucial proteins in collagen biosynthesis, matura...
Equine endometrial development during late fetal and postnatal periods. Equine uterine development, including endometrial histogenesis, begins prenatally and is completed postnatally. Little is known about this process in the horse. Uterine tissue was acquired from 38 foals, ranging in developmental age from gestational day (GD) 300 to postnatal day (PND) 180, for assessment of endometrial histogenesis. Patterns of endometrial cell proliferation were evaluated by multispectral imaging of uterine tissue sections stained immunofluorescently for Ki-67. Labeling index (LI, % labeled cells) for Ki-67 was calculated for each endometrial cell compartment (luminal epithel...
Age, gender and climate associations with the seroprevalence of Neospora species infection in horses in Jordan. A cross-sectional study was carried out on a sample of 379 horses to determine the seroprevalence of Neospora spp. in Jordan using the indirect fluorescent antibody test. Five variables, namely locality (n=10), climatic zone (n=4), age group (n=3), gender, and breed were tested as risk factors for Neospora-immunoglobulin (Ig)G seropositivity at four cutoff titers (1:50, 1:200, 1:400, and 1:800) using univariate and multivariate logistic regression analyses. A total of 122 (32%; 95% CI: 28, 37) sera samples had anti-Neospora-IgG at a cutoff titer of 1:50. Increased Neospora-IgG seropositivity w...
Quantitative Assessment of Tendon Hierarchical Structure by Combined Second Harmonic Generation and Immunofluorescence Microscopy. Histological evaluation of healing tendons is primarily focused on monitoring restoration of longitudinal collagen alignment, although the elastic property of energy-storing flexor tendons is largely attributed to interfascicular sliding facilitated by the interfascicular matrix (IFM). The objectives of this study were to explore the utility of second harmonic generation (SHG) imaging to objectively assess cross-sectional tendon fascicle architecture, to combine SHG microscopy with elastin immunofluorescence to assess the ultrastructure of collagen and elastin in longitudinal and transverse se...
Preparation of Monoclonal Antibody Against EMA-1 and Development of Rapid Serological Detection Method for Theileria equi Infection, Xinjiang, China. The erythrocytic-stage surface protein equi merozoite antigen 1 (EMA-1) of Theileria equi is a major candidate for the development of a diagnostic antigen for equine piroplasmosis. In this study, BALB/c mice were immunized with purified recombinant EMA-1 to prepare monoclonal antibody (mAb) against T. equi EMA-1, and 1 mAb 5H2 was obtained that showed good reaction with infected red blood cells (RBC) in the indirect immunofluorescence assay (IFA). To develop a rapid serological detection method for T. equi infection in Xinjiang Uygur Autonomous Region, China, recombinant EMA-1 originating from...
Phenotypic characterization of equine monocyte-derived dendritic cells generated ex vivo utilizing commercially available serum-free medium. The impact of culture conditions on equine monocyte-derived dendritic cells (MoDC) generation has not been fully characterized. We hypothesized that 1) MoDC could be cultured in a commercially available serum-free medium (AIM-V); and 2) that differential culture conditions would influence MoDC viability, yield and phenotype. MoDC generated from adult horses were cultured under variable conditions in a series of experiments. Viability was assessed using trypan blue and propidium iodide staining. Yield was determined by manual hemocytometer counting. Phenotype was assessed by flow cytometric ana...
Transplacental Transmission of Theileria equi Is Not a Common Cause of Abortions and Infection of Foals in Israel. Although the main route of transmission of is through tick feeding, transplacental transmission is also possible and may lead to abortion, or to the birth of a sick or carrier foal. The aim of this study was to evaluate the role of as a cause of abortions in Israel and the risk of foals being infected at a young age. Eight aborting mares were serologically evaluated for exposure to via the immunofluorescence antibody test (IFAT) and their aborted fetuses were evaluated using PCR and qPCR. In addition, five mares and their foals (aged 4-6 months) from a highly endemic farm were tested for i...
Genetic Characterization of Piroplasms in Donkeys and Horses from Nigeria. Equine piroplasmosis (EP) is a tick-borne disease of equids, caused by the two haemoprotozoal parasites: and Nigeria constitutes a major crossroads of animal transport in West Africa and may serve as a factor in EP dissemination in the region. The study aim was to characterize EP parasites in donkeys and horses in northern Nigeria using a molecular approach. Blood was collected from 57 donkeys and 47 horses. EP infection was detected and characterized by polymerase chain reaction (PCR). Twenty five donkeys (43.8%) were infected with , five (8.8%) with three (5.3%) with dual infections. Four...
Comparison of immunofluorescence and chemiluminescence assays for measuring ACTH in equine plasma. The chemiluminescence (CL) and immunofluorescence (IF) assays yield different results for basal adrenocorticotropin hormone concentrations [ACTH] in pony plasma. It is unclear whether this difference also occurs in basal samples from horses or samples from ponies following thyrotropin-releasing hormone (TRH) stimulation. Objective: To compare the results of [ACTH] analysis by CL and IF methods in basal samples from horses and pony samples following TRH stimulation. Methods: Method comparison. Methods: Plasma [ACTH] was measured concurrently using CL and IF methods in 12 ponies (basal and post-...
A Chimeric Sudan Virus-Like Particle Vaccine Candidate Produced by a Recombinant Baculovirus System Induces Specific Immune Responses in Mice and Horses. Ebola virus infections lead to severe hemorrhagic fevers in humans and nonhuman primates; and human fatality rates are as high as 67%-90%. Since the Ebola virus was discovered in 1976, the only available treatments have been medical support or the emergency administration of experimental drugs. The absence of licensed vaccines and drugs against the Ebola virus impedes the prevention of viral infection. In this study, we generated recombinant baculoviruses (rBV) expressing the Sudan virus (SUDV) matrix structural protein (VP40) (rBV-VP40-VP40) or the SUDV glycoprotein (GP) (rBV-GP-GP), and SUDV...
Neutrophil Extracellular Traps in the Pathogenesis of Equine Recurrent Uveitis (ERU). Equine recurrent uveitis (ERU) is considered one of the most important eye diseases in horses and typically appears with relapsing inflammatory episodes without systemic effects. Various disorders have been described as an initial trigger, including infections. Independent of the initiating cause, there are numerous indications that ERU is an immune-mediated disease. We investigated whether neutrophil extracellular traps (NETs) are part of the ERU pathogenesis. Therefore, vitreous body fluids (VBF), sera, and histological sections of the eye from ERU-diseased horses were analyzed for the prese...
