Preparation and characterization of a polyclonal antibody against brominated protein

Yasuhiro Kambayashi, Keiki Ogino, Kei Takemoto, Takashi Imagama, Tomoko Takigawa, Shingo Kimura, Yuri Hibino, Yoshiaki Hitomi, Hiroyuki Nakamura

Research output: Contribution to journalArticle

7 Citations (Scopus)

Abstract

(Di)bromotyrosine is formed by the specific reaction of eosinophil peroxidase and can be used as an eosinophil activation marker. In the present study, an antibody for (di)bromotyrosine in proteins was prepared to investigate the pathogenesis of eosinophil-related diseases such as allergic responses. A rabbit polyclonal antibody was raised against brominated keyhole limpet hemocyanin. The specificity of the antiserum was investigated with an enzyme-linked immunosorbent assay (ELISA). The antiserum recognized brominated bovine serum albumin (BSA) and dibromotyrosine-conjugated BSA. The antiserum also reacted with chlorinated BSA and di-iodotyrosine-conjugated BSA. Moreover, the specificity of the antiserum was investigated using competitive ELISA. Dibromotyrosine and di-iodotyrosine inhibited the recognition of brominated BSA by the antiserum. However, the recognition of brominated BSA by the antiserum was not inhibited by bromotyrosine, chlorotyrosine, iodotyrosine, nitrotyrosine, aminotyrosine, phosphotyrosine, or tyrosine. These results suggested that the epitope of the antiserum is dihalogenated tyrosine. Immunohistochemically, the antiserum stained brominated rat eosinophils but not chlorinated or nitrated eosinophils. In conclusion, an antiserum for dihalogenated protein was prepared. It is expected that the antiserum will be useful for the analysis of the pathogenesis of allergic diseases such as asthma and atopic dermatitis.

Original languageEnglish
Pages (from-to)95-103
Number of pages9
Journaljournal of clinical biochemistry and nutrition
Volume44
Issue number1
DOIs
Publication statusPublished - Jan 2009

Keywords

  • (Di)bromotyrosine
  • Allergic disease
  • Eosinophil activation marker
  • Oxidative stress
  • Polyclonal antibody

ASJC Scopus subject areas

  • Medicine (miscellaneous)
  • Nutrition and Dietetics
  • Clinical Biochemistry

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  • Cite this

    Kambayashi, Y., Ogino, K., Takemoto, K., Imagama, T., Takigawa, T., Kimura, S., Hibino, Y., Hitomi, Y., & Nakamura, H. (2009). Preparation and characterization of a polyclonal antibody against brominated protein. journal of clinical biochemistry and nutrition, 44(1), 95-103. https://doi.org/10.3164/jcbn.08-196