Immunological strategy for detecting the pro-inflammatory cytokine TNF-alpha in salmonids

Background: Tumour necrosis factor-alpha (TNF-α) is a pro-inflammatory cytokine which exerts a variety of immunological functions in vertebrates. TNF-α has been identified and cloned in a number of teleost fish species; nevertheless, the functions displayed by this cytokine in fish...

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Autores principales:	Rojas,Verónica, Morales-Lange,Byron, Guzmán,Fanny, Gallardo,José A, Mercado,Luis
Lenguaje:	English
Publicado:	Pontificia Universidad Católica de Valparaíso 2012
Materias:	fish immunity salmonid immunity TNF-alpha antibody
Acceso en línea:	http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0717-34582012000500021
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spelling	oai:scielo:S0717-345820120005000212012-11-29Immunological strategy for detecting the pro-inflammatory cytokine TNF-alpha in salmonidsRojas,VerónicaMorales-Lange,ByronGuzmán,FannyGallardo,José AMercado,Luis fish immunity salmonid immunity TNF-alpha antibody Background: Tumour necrosis factor-alpha (TNF-α) is a pro-inflammatory cytokine which exerts a variety of immunological functions in vertebrates. TNF-α has been identified and cloned in a number of teleost fish species; nevertheless, the functions displayed by this cytokine in fishes remain poorly understood, given that the low sequence identity compared to their mammalian counterpart, limit fish TNF-α detection using mammalian antibodies. Then, for fish immune response characterization is fundamental the production of specific fish anti-TNF-α antibody. Results: We have developed a monoespecific antibody against the pro-inflammatory molecule TNF-α of salmonid fish. TNF-α epitope region was identified and characterized using bioinformatic tools. The epitope sequence was chemically synthesized using Fmoc strategy, analyzed by RP-HPLC and its molecular weight confirmed by mass spectrometry. The synthetic peptide was used to immunize mice and antibodies from ascitic fluid were purified. The resulting antibody was used for molecular and histochemical detection in gut samples from salmonid fishes treated with different food. By ELISA, we detected a differential expression of TNF-α, the western blot analysis shows recognition of the whole TNF molecule and by immunohistochemistry TNF-α positive cells were observed. Conclusions: We provide an immunological tool, validated through classical immunological assays, which can be a useful tool for characterizing fish TNF-α function.info:eu-repo/semantics/openAccessPontificia Universidad Católica de ValparaísoElectronic Journal of Biotechnology v.15 n.5 20122012-09-01text/htmlhttp://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0717-34582012000500021en
institution	Scielo Chile
collection	Scielo Chile
language	English
topic	fish immunity salmonid immunity TNF-alpha antibody
spellingShingle	fish immunity salmonid immunity TNF-alpha antibody Rojas,Verónica Morales-Lange,Byron Guzmán,Fanny Gallardo,José A Mercado,Luis Immunological strategy for detecting the pro-inflammatory cytokine TNF-alpha in salmonids
description	Background: Tumour necrosis factor-alpha (TNF-α) is a pro-inflammatory cytokine which exerts a variety of immunological functions in vertebrates. TNF-α has been identified and cloned in a number of teleost fish species; nevertheless, the functions displayed by this cytokine in fishes remain poorly understood, given that the low sequence identity compared to their mammalian counterpart, limit fish TNF-α detection using mammalian antibodies. Then, for fish immune response characterization is fundamental the production of specific fish anti-TNF-α antibody. Results: We have developed a monoespecific antibody against the pro-inflammatory molecule TNF-α of salmonid fish. TNF-α epitope region was identified and characterized using bioinformatic tools. The epitope sequence was chemically synthesized using Fmoc strategy, analyzed by RP-HPLC and its molecular weight confirmed by mass spectrometry. The synthetic peptide was used to immunize mice and antibodies from ascitic fluid were purified. The resulting antibody was used for molecular and histochemical detection in gut samples from salmonid fishes treated with different food. By ELISA, we detected a differential expression of TNF-α, the western blot analysis shows recognition of the whole TNF molecule and by immunohistochemistry TNF-α positive cells were observed. Conclusions: We provide an immunological tool, validated through classical immunological assays, which can be a useful tool for characterizing fish TNF-α function.
author	Rojas,Verónica Morales-Lange,Byron Guzmán,Fanny Gallardo,José A Mercado,Luis
author_facet	Rojas,Verónica Morales-Lange,Byron Guzmán,Fanny Gallardo,José A Mercado,Luis
author_sort	Rojas,Verónica
title	Immunological strategy for detecting the pro-inflammatory cytokine TNF-alpha in salmonids
title_short	Immunological strategy for detecting the pro-inflammatory cytokine TNF-alpha in salmonids
title_full	Immunological strategy for detecting the pro-inflammatory cytokine TNF-alpha in salmonids
title_fullStr	Immunological strategy for detecting the pro-inflammatory cytokine TNF-alpha in salmonids
title_full_unstemmed	Immunological strategy for detecting the pro-inflammatory cytokine TNF-alpha in salmonids
title_sort	immunological strategy for detecting the pro-inflammatory cytokine tnf-alpha in salmonids
publisher	Pontificia Universidad Católica de Valparaíso
publishDate	2012
url	http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0717-34582012000500021
work_keys_str_mv	AT rojasveronica immunologicalstrategyfordetectingtheproinflammatorycytokinetnfalphainsalmonids AT moraleslangebyron immunologicalstrategyfordetectingtheproinflammatorycytokinetnfalphainsalmonids AT guzmanfanny immunologicalstrategyfordetectingtheproinflammatorycytokinetnfalphainsalmonids AT gallardojosea immunologicalstrategyfordetectingtheproinflammatorycytokinetnfalphainsalmonids AT mercadoluis immunologicalstrategyfordetectingtheproinflammatorycytokinetnfalphainsalmonids
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Immunological strategy for detecting the pro-inflammatory cytokine TNF-alpha in salmonids

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