Development of a rapid, high sensitive, simple method for the determination of endocrine disrupters in environment using solid phase extraction

将抗体固定的微球转移到JR细胞中，建立了鲤鱼卵黄蛋白原(VG)的顺序注射分析(SIA)技术。这一测定是基于夹心免疫分析，其中使用了抗鲤鱼VG抗体和鲤鱼VG之间的两种反应。也就是说，第一步反应的抗体固定在微球(Sephadex珠子)上，第二步反应中使用辣根过氧化物酶(HRP)标记的抗体。反应中用过氧化氢和邻苯二胺(OPD)的混合溶液作为发色团的来源。将微球固定的抗体、VG分析物、HRP标记的抗体和显色液按程序自动引入SIA系统的JR细胞，并利用氧化OPD产物的吸光度来确定VG的存在数量。考虑到对VG测定的敏感性，第一步和第二步免疫反应的最佳孵育时间分别为120分钟和60分钟。在此条件下，VG浓度与氧化OPD的吸光度之间具有良好的线性关系。该体系测定VG的检出限为5ngml1左右。本方法是一种简便、准确的VG测定方法。

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 antibody 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 immunoreaction 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.