卵黄抗体用于眼镜王蛇毒抗原检测的初步研究

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卵黄抗体用于眼镜王蛇毒抗原检测的初步研究

王桂平, 刘新艳, 朱柳, 覃媛, 黄劭, 余清声^*

1.广州医学院药物研究中心, 广东广州, 510182
2.广州医学院护理学院, 广东广州, 510182;
3.广州医学院广州蛇毒研究所, 广东广州, 510182

收稿日期:2005-02-04 修回日期:2005-08-10 出版日期:2005-09-15 发布日期:2005-09-15
通讯作者: 余清声*

Primary Study of Egg Yolk Antibody for Detection of king cobra Venom Antigens

WANG Gui-ping, LIU Xin-yan, ZHU Liu, QIN Yuan, HUANG Shao, YU Qing-sheng^*

1.Drug Research Center, Guangzhou Medical College, Guangzhou;
2 Nursing School, Guangzhou Medical College;
3 Guangzhou Research Institute of Snake Venom, Guangzhou Medical College, 510182, China

Received:2005-02-04 Revised:2005-08-10 Online:2005-09-15 Published:2005-09-15
Contact: YU Qing-sheng*

摘要/Abstract

摘要：

目的探讨抗蛇毒卵黄抗体应用于蛇毒检测的可行性。方法以甲醛减毒的眼镜王蛇毒抗原免疫莱航母鸡, 制备抗眼镜王蛇毒抗体; 采用过碘酸钠法对特异卵黄抗体进行酶标; ELISA的实验条件及相关参数通过棋盘滴定法确定, 并进行特异性、灵敏度、精确度及稳定性等测定。结果本检测方法的灵敏度可达32 μg·L^-1, 并且眼镜王蛇毒抗原浓度在32～750 μg·L^-1的范围内, 线性关系较好(r = 0.963); 卵黄抗体对眼镜王蛇毒具有较高的特异性, 与蝰蛇科的五步蛇毒、园斑蝰蛇毒无交叉反应, 与眼镜蛇科的金环及银环蛇毒仅在高剂量(剂量大于500 μg·L^-1)时存在轻度交叉反应, 但与眼镜蛇毒之间交叉反应明显。应用本方法对不同浓度(100、300、1000 μg·L^-1)的眼镜王蛇毒样品检测, 结果平均板内相对标准差在1%～3%之内, 板间相对标准差在8%以内; 稳定性实验显示:卵黄抗体置于37 ºC条件6天, 其检测结果与4 ºC条件下无显著差异(P>0.05)。结论特异的卵黄抗体应用于微量蛇毒检测研究是可行的, 但仍需要进一步研究以降低交叉反应。

关键词: 眼镜王蛇毒, 眼镜王蛇毒, 眼镜王蛇毒, 卵黄抗体IgY, 卵黄抗体IgY, 卵黄抗体IgY, 母鸡, 母鸡, 母鸡, 酶联免疫吸附测定ELISA, 酶联免疫吸附测定ELISA, 酶联免疫吸附测定ELISA

Abstract: Aim To study whether antivenom from laying hens can be used for the detection of venom antigens. Methods Chickens (white Leghorn) were immunized with detoxicated king cobra venom by formaldehyde and egg yolk immunoglobulin (IgY) isolated from yolk; IgY was labelled with the horseradish peroxidase (HRP). Experimental condition and parameters were determined by chessboard test. The specificity, sensitivity, precision, and stability of this method were assayed in the experiment. Results This method could detect as low as 32 μg·L^-1 of the king cobra antigens. A good linear relation was found within 32 - 750 μg·L^-1of king cobra venom concentrations (r = 0.963). There was no cross reactivity for the reagents with Agkistrodon acutus Guenther venom or Vipera russelli siamensis Smith venom; slight cross reactivity with Bungarus multicinctus Blyth venom or Bungarus fasciatus Chmeider venom; and notable cross reactivity with cobra venom. The average intra-assay relative standard deviation (RSD) was 1%-3%, and the inter-assay RSD was less than 8%. The reagents (including IgY and HRP-IgY) were stable; no differences (P>0.05) were observed for the detection of venom antigens when the reagents were stored at 37 ºC up to 6 d. Conclusion IgY is a good reagent for diagnosis of snakebite after eliminating the genus cross reactivity.

Key words: king cobra venom, king cobra venom, yolk immunoglobulin, yolk immunoglobulin, hens, hens, enzyme-linked immunosorbent assays, enzyme-linked immunosorbent assays

中图分类号:

R991

R996.3

Supporting: Foundation item: Supported by Guangzhou Technological Bureau (No:2002J1-C0l7l).
^*Corresponding author. Tel.: 020-81340243

王桂平, 刘新艳, 朱柳, 覃媛, 黄劭, 余清声^*. 卵黄抗体用于眼镜王蛇毒抗原检测的初步研究[J]. .

WANG Gui-ping, LIU Xin-yan, ZHU Liu, QIN Yuan, HUANG Shao, YU Qing-sheng^*. Primary Study of Egg Yolk Antibody for Detection of king cobra Venom Antigens[J]. .

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