Spectrophotometric determination of carp vitellogenin using a sequential injection analysis technique equipped with a jet ring cell

Nobuaki Soh, Hideshi Nishiyama, Keiko Mishima, Toshihiko Imato, Takashi Masadome, Yasukazu Asano, Youichi Kurokawa, Hisao Tabei, Saeko Okutani

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

A sequential injection analysis (SIA) technique, in which antibody-immobilized microbeads were transferred to a jet ring (JR) cell, was used in determination of carp vitellogenin (Vg). The determination is based on a sandwich immunoassay in which two types of reactions between anti carp Vg antibodies and carp Vg are used. Namely, the antibody for the first reaction step was immobilized on microbeads (Sephadex beads), and an antibody labeled with a horseradish peroxidase (HRP) was used in the second step of the reaction. A mixed solution of hydrogen peroxide and o-phenylenediamine (OPD) was used as the source of the chromophore in the reaction. The microbeads-immobilized antibody, Vg analyte, HRP-labeled anitbody and the color developing solution were introduced automatically into the JR cell of the SIA system in a programmed sequence, and the absorbance of the oxidized OPD product was used to determine the amount of Vg present. The optimal incubation times for the immuno-raction for the first and the second steps were determined at 120 and 60 min, respectively, taking into account the sensitivity to the Vg determination. Under these conditions, a good linear correlation was obtained between Vg concentration and the absorbance of the oxidized OPD. The lower detection limit for the determination of Vg was about 5 ng ml-1 in this system. The method developed here represents a simple, accurate method for the determination method of Vg.

Original languageEnglish
Pages (from-to)1123-1130
Number of pages8
JournalTalanta
Volume58
Issue number6
DOIs
Publication statusPublished - 2002 Dec 6
Externally publishedYes

Keywords

  • Antigen-antibody reaction
  • Beads suspension
  • Sequential injection
  • Spectrophotometry
  • Vitellogenin

ASJC Scopus subject areas

  • Analytical Chemistry

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