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13 pages, 1411 KiB

Open AccessArticle

Enzyme-Linked Immunosorbent Assay (ELISA) Development for Equine Serum Amyloid A (SAA) Determination Using Recombinant Proteins

by Pollyanna C. Souto, Marcus R. Santos, Andrés M. Ortega Orozco, Lucas D. Bento, Camilo J. Ramirez-Lopez, Fabrícia M. Girardi, Júlia C. Assis Machado, Leandro L. de Oliveira and Leandro A. da Fonseca

Methods Protoc. 2025, 8(2), 37; https://doi.org/10.3390/mps8020037 - 7 Apr 2025

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Abstract

We aimed to develop a species-specific ELISA for qualitatively and quantitatively determining serum amyloid A (SAA) in horses. Current methods for measuring SAA in horses utilize ELISA or immunoturbidimetric tests designed for human SAA, which are not specific to horses. Mice and rabbits were used to generate polyclonal antibodies against equine SAA. The study examined serum samples from 32 horses with acute inflammatory disease (SG) and 25 clinically healthy horses. Furthermore, the SAAeq kinetics were observed in three horses from the SG group at three different timepoints. The SAA-ELISA established a cut-off at 0.06 OD₄₉₂nm, where values equal to or higher than this were deemed positive, while values below it was considered negative. The test exhibited a sensitivity of 94% and specificity of 92%, resulting in an overall accuracy of 93%. The positive and negative predictive values were 94% and 92%, respectively. Coefficients of variation for inter- and intra-assay were 6.1% and 7.46% for SG and 9.6% and 9.63% for the control group (CG). The detection limit was determined to be 0.067. The SAA-ELISA proved its worth by demonstrating satisfactory performance, paving the way for the development of automated quantitative tests and species-specific semi-quantitative tests. This paves the way for their application in practical field settings. Full article

(This article belongs to the Section Molecular and Cellular Biology)

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15 pages, 5093 KiB

Open AccessArticle

Dynamic Correlation between Platelet Aggregation and Inflammatory-like State in Athlete Horses

by Francesca Arfuso, Maria Rizzo, Federica Arrigo, Caterina Faggio, Elisabetta Giudice, Giuseppe Piccione and Claudia Giannetto

Appl. Sci. 2024, 14(5), 2086; https://doi.org/10.3390/app14052086 - 1 Mar 2024

Cited by 5 | Viewed by 1355

Abstract

This study aimed to assess the effect of exercise on serum electrophoretic protein pattern, C-reactive protein (CRP) and platelet aggregation in horses subjected to a jumping exercise. The possible relationship between acute-phase reactions and platelet reactivity in the context of exercise was investigated. Blood samples were collected from 10 jumper horses at rest (TREST), within 5 min from the end of exercise (TPE5), and 30 min (TPE30) and 60 min after exercise (TPE60). The serum values of total proteins; CRP; albumin; α1-, α-2, β1-, β2- and γ-globulins; and the maximum degree of aggregation and the initial velocity of aggregation (slope) were evaluated. According to one-way analysis of variance, CRP and α1-, α-2, β1- and β2-globulins increased after exercise compared with rest condition (p < 0.001), whereas albumin and platelet aggregation showed lower values after exercise than at rest (p < 0.001). CRP and α1-globulin values were negatively correlated with both platelet aggregation indices at TPE5, whereas no significant correlation among these parameters was found at TREST, TPE30 and TPE60. This study provides evidence that an acute-phase response occurred in horses after the jumping exercise and suggests a linkage between the inflammatory status and the platelet responsiveness in horses during exercise. Full article

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17 pages, 345 KiB

Open AccessReview

Biomarkers of Intestinal Injury in Colic

by Elsa K. Ludwig, Kallie J. Hobbs, Caroline A. McKinney-Aguirre and Liara M. Gonzalez

Animals 2023, 13(2), 227; https://doi.org/10.3390/ani13020227 - 7 Jan 2023

Cited by 11 | Viewed by 3956

Abstract

Biomarkers are typically proteins, enzymes, or other molecular changes that are elevated or decreased in body fluids during the course of inflammation or disease. Biomarkers pose an extremely attractive tool for establishing diagnoses and prognoses of equine gastrointestinal colic, one of the most prevalent causes of morbidity and mortality in horses. This topic has received increasing attention because early diagnosis of some forms of severe colic, such as intestinal ischemia, would create opportunities for rapid interventions that would likely improve case outcomes. This review explores biomarkers currently used in equine medicine for colic, including acute phase proteins, proinflammatory cytokines, markers of endotoxemia, and tissue injury metabolites. To date, no single biomarker has been identified that is perfectly sensitive and specific for intestinal ischemia; however, L-lactate has been proven to be a very functional and highly utilized diagnostic tool. However, further exploration of other biomarkers discussed in this review may provide the key to accelerated identification, intervention, and better outcomes for horses suffering from severe colic. Full article

(This article belongs to the Special Issue Equine Colic)

28 pages, 4490 KiB

Open AccessEditor’s ChoiceReview

IgG Fusion Proteins for Brain Delivery of Biologics via Blood–Brain Barrier Receptor-Mediated Transport

by Ruben J. Boado

Pharmaceutics 2022, 14(7), 1476; https://doi.org/10.3390/pharmaceutics14071476 - 15 Jul 2022

Cited by 19 | Viewed by 5570

Abstract

The treatment of neurological disorders with large-molecule biotherapeutics requires that the therapeutic drug be transported across the blood–brain barrier (BBB). However, recombinant biotherapeutics, such as neurotrophins, enzymes, decoy receptors, and monoclonal antibodies (MAb), do not cross the BBB. These biotherapeutics can be re-engineered as brain-penetrating bifunctional IgG fusion proteins. These recombinant proteins comprise two domains, the transport domain and the therapeutic domain, respectively. The transport domain is an MAb that acts as a molecular Trojan horse by targeting a BBB-specific endogenous receptor that induces receptor-mediated transcytosis into the brain, such as the human insulin receptor (HIR) or the transferrin receptor (TfR). The therapeutic domain of the IgG fusion protein exerts its pharmacological effect in the brain once across the BBB. A generation of bifunctional IgG fusion proteins has been engineered using genetically engineered MAbs directed to either the BBB HIR or TfR as the transport domain. These IgG fusion proteins were validated in animal models of lysosomal storage disorders; acute brain conditions, such as stroke; or chronic neurodegeneration, such as Parkinson’s disease and Alzheimer’s disease. Human phase I–III clinical trials were also completed for Hurler MPSI and Hunter MPSII using brain-penetrating IgG-iduronidase and -iduronate-2-sulfatase fusion protein, respectively. Full article

(This article belongs to the Special Issue Advanced Blood-Brain Barrier Drug Delivery)

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14 pages, 1375 KiB

Open AccessArticle

Usefulness of Selected Acute-Phase Proteins in the Postsurgical Monitoring of Arthroscopy and Splint Bone Removal in Horses

by Maciej Przewoźny, Magdalena Senderska-Płonowska, Anna Rząsa, Heliodor Wierzbicki, Jacek Borkowski, Jan-Hein Swagemakers, Agnieszka Żak-Bochenek and Tadeusz Stefaniak

Animals 2021, 11(10), 2952; https://doi.org/10.3390/ani11102952 - 13 Oct 2021

Viewed by 2061

Abstract

Background: Arthroscopy and splint bone removal are the common orthopedic procedures in horses. Estimation of the dynamics of acute phase proteins in postoperative monitoring seems to be interesting diagnostic approach. The aim of the study was to investigate changes in the concentrations of plasma inflammatory markers—fibrinogen, haptoglobin, and protease inhibitors—following orthopedic surgery in horses. The study involved 114 horses, divided into two study groups undergoing: arthroscopy (41 horses) and splint bone removal (13 horses). The control group consisted of 60 healthy horses. The blood was collected before the surgery and 24, 48, 72 h, 5, 7, 10, 14 and 28 days after the surgery. Plasma fibrinogen, serum haptoglobin and proteinase inhibitors were measured. Results: In non-complicated cases of arthroscopy and splint bone removal, fibrinogen and haptoglobin increased stepwise from 24 h, achieved the maximum level at 72 h and returned to preoperative levels after 10–14 days. In one complicated case after arthroscopy surgery the marked increase in fibrinogen and haptoglobin concentrations was observed 24 h earlier than standard parameters of inflammation Conclusion: The study shows the evolution of APPs after arthroscopy and splint bone removal in 28 days postsurgery period and in the case of one complicated case of arthroscopy. Full article

(This article belongs to the Section Veterinary Clinical Studies)

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