Establishment and application of double-antibody sandwich ELISA for determination of 3-nitrotyrosine / 中国应用生理学杂志
Chinese Journal of Applied Physiology
; (6): 569-572, 2009.
Article
en Zh
| WPRIM
| ID: wpr-301559
Biblioteca responsable:
WPRO
ABSTRACT
<p><b>AIM</b>To prepare the working standards of 3-nitrotyrosine (3-NT) and establish a two-antibody-sandwich ELISA for determining the concentration of peroxynitrite in the tissue.</p><p><b>METHODS</b>Nitrated bovine serum albumin was prepared by additions of an alkaline stock solution of peroxynitrite which was synthesized by a quenched-flow reactor. The monoclone anti-3-NT antibody from mouse was used as coating antibody and the polyclone anti-3-NT antibody from as labeling antibody to prepare the standard work curve by orthogonal design. The concentrations of 3-NT in cardiac tissue from rats subjected to myocardial ischemia and reperfusion (MI/R) were analyzed.</p><p><b>RESULTS</b>A two-antibody-sandwich ELISA method for measuring 3-NT content in biological fluids and homogenates was successfully established. The detecting limit was 0.1 ng x ml(-1) and the linear range of standard work curve was 0.15 - 7.50 ng x ml(-1) (r2 = 0.995). The 3-NT concentration in cardiac tissue from rats subjected to MI/R (1022.42 +/- 97.35 ng x mg pro(-1)) was significantly higher than that in the sham group (246.58 +/- 56.52 ng x mg pro(-1), P < 0.01).</p><p><b>CONCLUSION</b>A two-antibody-sandwich ELISA was established for determining the 3-NT concentration in the tissue and conveniently, quickly, accurately quantitative analysis of the content of 3-NT. The assay provides a new method for quantitative analysis of the peroxyinitrite in the future.</p>
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1
Índice:
WPRIM
Asunto principal:
Tirosina
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Ensayo de Inmunoadsorción Enzimática
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Daño por Reperfusión Miocárdica
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Química
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Sensibilidad y Especificidad
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Ratas Wistar
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Ácido Peroxinitroso
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Métodos
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Anticuerpos Monoclonales
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Miocardio
Tipo de estudio:
Diagnostic_studies
Límite:
Animals
Idioma:
Zh
Revista:
Chinese Journal of Applied Physiology
Año:
2009
Tipo del documento:
Article