Methodological aspects of assessing phagocytosis of Vibrio anguillarum by leucocytes of gilthead seabream (Sparus aurata L.) by flow cytometry and electron microscopy

Cell and Tissue Research
M A EstebanJ Meseguer

Abstract

In this paper we optimize a flow cytometric method for evaluating the phagocytic activity of leucocytes in gilthead seabream (Sparus aurata L.) and characterize the phagocytic cells observed. Optimal conditions were established for the fluorescein-labelling and analysis of the bacterium Vibrio anguillarum by flow cytometry. Head-kidney leucocytes were incubated with the heat-killed fluorescein isothiocyanate (FITC)-labelled bacteria for different periods, during which the kinetics of phagocytosis was studied. Attached and interiorized bacteria were distinguished. Although phagocytic ability reached a maximum after 60 min, phagocytic capacity reached its maximum at 20 min. The amount of ingested bacteria per phagocyte was estimated from the mean fluorescence of the leucocytes. Cytochalasin B or colchicine was used to inhibit phagocytosis. Monocyte-macrophages and acidophilic granulocytes showed phagocytic activity as demonstrated by transmission electron microscopy. In conclusion, the technique presented allows the screening of thousands of cells, and individual cell evaluation, by quantifying interiorized particles in fish phagocytes. Our ultrastructural results demonstrate that V. anguillarum is actively phagocytized by seabre...Continue Reading

Citations

Jun 9, 2012·Fish Physiology and Biochemistry·Rebeca CerezuelaM Ángeles Esteban
Jun 25, 2005·Developmental and Comparative Immunology·Irene SalinasJosé Meseguer
Jul 27, 2001·Developmental and Comparative Immunology·V MuleroJ Meseguer
Dec 13, 2002·Developmental and Comparative Immunology·Alberto CuestaJosé Meseguer
Sep 5, 2002·Veterinary Immunology and Immunopathology·Jesús OrtuñoJ Meseguer
Dec 22, 1998·Veterinary Immunology and Immunopathology·V MuleroJ Meseguer
Jun 4, 2014·Developmental and Comparative Immunology·Erick García-GarcíaVictoriano Mulero
Dec 3, 2003·Developmental and Comparative Immunology·Pablo PelegrínJosé Meseguer
Apr 22, 2003·The Anatomical Record. Part A, Discoveries in Molecular, Cellular, and Evolutionary Biology·Alejandro RodríguezJosé Meseguer
Oct 25, 2011·Developmental and Comparative Immunology·Isabel CabasAlfonsa García-Ayala
Jul 20, 2011·Fish & Shellfish Immunology·Martha Reyes-BecerrilMaría Ángeles Esteban
Dec 21, 2010·Fish & Shellfish Immunology·F A GuardiolaM A Esteban
Aug 10, 2010·Developmental and Comparative Immunology·S LiarteA García-Ayala
Dec 9, 2008·Fish & Shellfish Immunology·Rebeca CerezuelaMa Angeles Esteban
Sep 28, 2015·Toxicology in Vitro : an International Journal Published in Association with BIBRA·Patricia MorcilloAlberto Cuesta
Oct 16, 2008·Fish & Shellfish Immunology·Ing-Fong JiangChing-Feng Weng
May 25, 2007·Fish & Shellfish Immunology·C DoñateS MacKenzie

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