鸡抗人精子卵黄抗体IgY的制备

目的 :探索一种快速和经济的从鸡卵黄制备抗人精子抗体IgY的方法。　方法 :用洗涤过的人精子免疫开产前的实验母鸡 ,间隔 7d加强免疫 ,4次加强注射后收集免疫鸡蛋 ,分离蛋黄和蛋清 ,去除蛋黄中的脂质后经 4次硫酸铵沉淀去除其它杂蛋白 ,再经SephadexG 2 5层析柱脱盐后获得卵黄抗体IgY。　结果 :鸡抗人类精子卵黄抗体IgY能使人类精子凝集 ,对照组用未免疫的鸡血清和卵黄抗体均为阴性。　结论 :产蛋母鸡可用来低成本和高效地生产抗人类精子抗体 ,精子凝集试验结果提示抗精子卵黄抗体可成为未来避孕药具的极有潜力的候选方法

Preparation of Antibodies against Human Sperm from Egg Yolks of Immunized Hens

WT5”HX]Objectives:[WT5”BZ]To develop a rapid and economical procedure for preparation and extraction of antibodies to human sperm from egg yolks of immunized hens. [WT5”HX]Mehtods:[WT5”BZ]Laying hens, 23 weeks old, were regularly administered by intramuscular injection with washed human sperm at an one week interval. Immunized eggs were collected after 4 booster injections for isolation of yolk immunoglobulins. The eggs were pooled, and the extraction procedure was run once a week. The yolk was separated from the white and carefully washed with Tris buffered saline (TBS) (1 mmol/L Tris, 14 mmol/L NaCl, pH7.4) to remove as much of the albumin as possible. The yolk membrane was cut and the yolk poured into a glass beaker and diluted with TBS. For one yolk (15ml), 40ml of TBS were added. The mixture was homogenized for ten min with a mixer. Then 40 ml of chloroform were added with continuous stirring. The mixture was then refrigerated overnight. Supernantant was transferred into a glass beaker and brought to room temperature. Half this volume of saturated ammonium sulphate solution was added to the supernantant to precipitate immunoglobulins. The precipitate was dissolved in a convenient volume of TBS. The re crystallization procedure was repeated four times to remove as much of the impure proteins as possible. The final precipitation was dissovled in a minimal volume of TBS. The extract was passed through a Sephodex G 25 column to remove sulphate. Barium chloride was used to trace the presence of sulphate in the extract. [WT5”HX]Results:[WT5”BZ]The presence of anti human sperm antibodies in the extracts was confirmed by sperm agglutination test. The control test with non immunized chicken IgY and chicken sera showed negative results in sperm agglutination test. [WT5”HX]Conclusions:[WT5”BZ]The phylogenetic distance between birds and human, the ability of chickens to produce high levels of specific antibodies following immunization with human sperm, and the transfer of large amounts of these antibodies from the serum of the laying hen to the yolk of the unfertilized egg have led to the development of immunoglobulins based passive immunocontraceptives in the near future. [WT5”HZ]