Browsing by Subject "Western blot"

Now showing 1 - 4 of 4
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    An optimized xylene-free protein extraction method adapted to formalin-fixed paraffin embedded tissue sections for western blot analysis
    (Universidad de Murcia. Departamento de Biología Celular e Histología, 2017) Mansour, Anthony G.; Abou Khalil, Pamela; Bejjani, Noha; Chatila, Rajaa; Dagher Hamalian, Carole; Faour, Wissam H.
    Deparaffinization of formalin-fixed paraffin embedded (FFPE) tissues with xylene currently remains a major challenge to the biomedical community. We developed an efficient xylene-free protocol to isolate proteins from archived FFPE human tissue sections. A total of 79 different types of FFPE tissue sections of 8 µm thickness were obtained from various archived FFPE specimens. Deparaffinization was conducted by gently washing each section with around 1 ml of hot distilled water (≈80°C). The deparaffinized tissues were homogenized in lysis buffer, and the isolated proteins were quantified and efficiently resolved using western blot analysis for the presence of Protein kinase B (PKB/AKT) and β-actin. Moreover, a significant amount of proteins was successfully isolated with an average of 2.31 µg/µl. The migration pattern of AKT and β-actin obtained from the specimens was similar to the positive control obtained from protein lysates prepared from in vitro cultured MDA231 cancer cell lines. AKT was successfully identified in all specimens, and β-actin protein was resolved with an efficiency higher than 80%. The entire extraction procedure requires only 20 minutes. This newly developed technique is an efficient, safe, cost-effective, and rapid method to isolate proteins from FFPE tissue sections adequate for molecular analysis.
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    Ezrin and moesin expression in canine and feline osteosarcoma
    (Universidad de Murcia. Departamento de Biología Celular e Histología, 2017) Hlavaty, Juraj; Wolfesberger, Birgitt; Hauck, Marlene; Obermayer Pietsch, Barbara; Fuchs Baumgartinger, Andrea; Miller, Ingrid; Walter, Ingrid
    Biological features of canine osteosarcomas (OS) differ markedly from those found in feline and resemble more human osteosarcomas, in particular for their high rate of metastasis and poor prognosis. Ezrin, radixin and moesin are members of the ERM protein family and link the actin cytoskeleton with the cell membrane. Ezrin and moesin have been shown to be of prognostic significance in tumor progression due to their role in the metastatic process. The objective of this study was to analyze ezrin and moesin protein expression in a series of dog (n=16) and cat (n=8) osteosarcoma samples using immunohistochemistry and western blot techniques. We found that cat OS have a higher moesin expression compared to dog OS, however, the active phosphorylated forms of moesin and ezrin Tyr353 were more abundant in the dog samples. A statistically significant difference was found for the low and high immunohistochemical scores of ezrin and pan-phosphoERM proteins between cat and dog. Although phosphoezrin Thr567 was higher in feline OS, the membranous localization in dog OS samples indicates the presence of the biologically active form. Therefore, the observed differences in phosphorylated forms of ezrin and moesin status should be further studied to demonstrate if they are relevant for different biological behavior between dog and cat OS.
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    Measurement of salivary adiponectin concentrations in dogs
    (Wiley , 2014-07-10) Tvarijonaviciute, Asta; Carrillo Sánchez, J. D.; García Martínez, Juan Diego; Tecles Vicente, Fernando; Martínez Subiela, Silvia; German, Alexander J.; Cerón Madrigal, José Joaquín; Medicina y Cirugía Animal; Facultades de la UMU::Facultad de Veterinaria
    Antecedentes: La medición de adiponectina salival podría mejorar la comprensión de la fisiología de esta adipocina y de su papel en diversas condiciones clínicas. Objetivos: El propósito del estudio fue evaluar la utilidad de un kit ELISA humano para adiponectina en la medición de adiponectina salival en perros, comparar las concentraciones de adiponectina en suero y saliva en una población de perros sanos, y evaluar los posibles efectos de la limpieza dental sobre las concentraciones de adiponectina sérica y salival en perros. Métodos: Para la validación analítica, se determinaron la precisión, exactitud y el límite inferior de cuantificación del ensayo utilizando muestras de saliva. Además, se cuantificaron las concentraciones de adiponectina en muestras de suero y saliva de 24 perros sanos, y de 7 perros con gingivitis leve antes y después de un procedimiento de limpieza dental. Resultados: Los ensayos de validación para adiponectina salival presentaron coeficientes de variación inferiores al 15%, y la recuperación osciló entre el 85% y el 120%. En la prueba de linealidad se observó interferencia al medir adiponectina en saliva, pero esto se resolvió diluyendo las muestras 1:4. En perros sanos, las concentraciones de adiponectina salival y sérica se correlacionaron positivamente (r = 0,650; P = 0,009). Tras el procedimiento de limpieza dental, la concentración de adiponectina salival aumentó el día 0 (P = 0,004), pero a los 14 días las concentraciones fueron inferiores a las registradas antes del procedimiento (P = 0,041). Conclusiones: El kit ELISA humano para adiponectina puede utilizarse para medir con precisión y exactitud la adiponectina salival en perros. La adiponectina salival aumentó 24 horas después de la limpieza dental, posiblemente debido a un proceso inflamatorio agudo o a la filtración de adiponectina desde la sangre tras el traumatismo gingival.
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    Seronegativity of Iberian ibex (Capra pyrenaica) against Toxoplasma gondii and Neospora caninum is consistent with eco-epidemiological and environmental features in Mediterranean mountainous areas
    (Elsevier, 2025-01-07) Cano Manuel, Alejandro; Granados, José Enrique; Álvarez García, Gema; Huertas López, Ana; Diezma Díaz, Carlos; Cano Manuel, Francisco Javier; Ortega Mora, Luis Miguel; Fandos, Paulino; Mentaberre, Gregorio; López Olvera, Jorge Ramón; Martínez-Carrasco Pleite, Carlos; Sanidad Animal; Facultades de la UMU::Facultad de Veterinaria