Veterinary Biosciences - Theses
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ItemD-Dimer in dogs, cats and horses: Investigations into immunoreactivity and structure( 2023)Background: D-dimer is a degradation product of cross-linked fibrin generated during thrombolysis. D-dimer is measured in human medicine for the early detection and exclusion of thrombotic syndromes. Commercially available D-dimer assays use monoclonal antibodies against human D-dimer, and there is limited sensitivity and specificity data for these antibodies in companion animals, thus the accuracy of human assays in veterinary species remains uncertain. Little is known about the sequence and structure of D-dimer in companion animals. Objectives: To evaluate the immunoreactivity of D-dimer in dogs, horses and cats with commercially available antibodies, and investigate potential differences in sequence and structure between species. Methods: A cross-linked fibrin lysate was prepared from canine, feline and equine plasma samples, and immunoblotting performed with a variety of commercially available D-dimer antibodies. Amino acid sequences of fibrinogen fragments involved in D-dimer were compared between species (cat, horse, dog and human), using available sequences for fibrinogen chains, and these were used for multiple sequence alignment, estimation of physiochemical properties, analysis of conserved structural domains, and prediction of three-dimensional structure by homology modelling. Results: The antibodies evaluated demonstrated variable reactivity with D-dimer in each species. The monoclonal antibody DD44 bound canine D-dimer with good specificity and sensitivity, but this antibody did not react with feline or equine D-dimer. The polyclonal antibody D2D bound putative D-dimer in dogs, cats and horses with good specificity, and increased sensitivity compared to human D-dimer. Analysis of sequence and structure revealed high inter-species homology, and three-dimensional models were prepared using a human template. Conclusions: Commercially available antibodies to human D-dimer have variable immunoreactivity with D-dimer in companion animals, suggesting that the use of human assays is inappropriate until analytical and/or clinical validation of the antibody used in that assay has been performed for each species. There are no overt structural variations to explain the variation in immunoreactivity between species, and this could reflect minor variations in structure at the site of binding for each antibody, which is largely unknown.