IgY抗体在免疫检测中的应用进展

卵黄免疫球蛋白(Immunoglobulin of yolk,IgY)是鸟类、爬行动物和两栖类动物体内的主要抗体,与哺乳动物IgG功能类似,一般从被抗原免疫的母鸡卵黄中提取。与IgG相比,IgY具有许多突出的优点,在制备针对多种抗原的多克隆抗体、免疫检测、人类和动物疾病的预防治疗等方面均得到广泛应用。对IgY在免疫检测方面的应用进展进行了综述。     

Research Progress on Immunoglobulin of Yolk in the Prevention and Treatment of Various Diseases

Immunoglobulin of yolk (IgY) refers to the specific antibodies from the immune eggs, is the most important immunoglobulin in the yolk,which has the characteristics of stable chemical properties, high yield, low cost, high specificity and no toxic side effects, there was no cross serological reaction among animals breeds because of a long distance. IgY has a significant advantage in animal and human disease prevention and treatment. In this research,we reviewed the application of structural characteristics, physicochemical property and action mechanism of IgY in prevention treatment of animal disease and developing prospect, especially in the treatment of animal diseases, such as diarrhea and acute treatment of animal spirit, harm, large virus disease; zoonosis seriously endanger the safety of human property; Other aspects such as aquaculture, animal fur and other major diseases. IgY could effectively prevent and treat animal diseases, but also reduce the possibility of zoonosis, which will be of great significance for the protection of personal safety.     

卵黄免疫球蛋白配基亲和纯化碱性磷酸酶

 以卵黄免疫球蛋白(immunoglobulin　yolk,IgY)为配基,对小牛肠组织来源的碱性磷酸酶进行免疫亲和纯化。采用1.5mol·L-1的NaCl进行洗脱,可使酶的纯化倍数达到64倍,活力回收率达到90%。IgY抗体对碱性磷酸酶亲和力的测定结果显示固有亲和力和相对亲和力分别为6.275×10-7　mol·L-1和5.0 μg·ml-1。     

五龙鹅IgY、IgM分子特性研究

为丰富和完善鹅Ig分子特性的研究,以层析、离子交换等方法分离纯化五龙鹅的免疫球蛋白及重链、轻链、Fab段、Fc段,进行鹅Ig的多态性、分子质量、氨基酸组成、稳定性等分子特性研究。结果表明:五龙鹅IgY存在几个亚类,总分子质量为166ku,重链约为60ku,轻链约为23.5ku;鹅IgM的重链分子质量分别为51.72、48.30和44.09ku,轻链20.30ku,所以,总分子质量约为660~740ku;五龙鹅Ig重链的分子质量均小于鸡、鸭;IgY的Fab段分子质量为48.67ku,Fc段为68.62ku。IgY的重链和轻链中碱性氨基酸摩尔分数小于酸性氨基酸,为酸性蛋白质,而IgM则为碱性蛋白质;IgY中Cys摩尔分数为2.25%,IgM为3.26%;鹅IgY的重链和轻链中Pro的摩尔分数相差较大,分别为7.69%和0.10%;鹅IgY在pH3.5~11.5的范围内十分稳定,IgM在pH4.0~10.5的范围内稳定,IgM的酸碱稳定性小于IgY,但在pH3.5~4.5时活性增加5%~8%;研究表明五龙鹅IgY热稳定性较差,热变性反应级数为1.2。     

鸡新城疫卵黄抗体IgY的分离提纯研究

本文对母鸡经鸡新城疫疫苗免疫接种后产生并富集在卵黄中的抗体IgY的提取纯化工艺条件进行了研究与优化，确定了去脂、提取抗体最佳工艺为：卵黄液用0．05mol／L,pH5.0的乙酸一乙酸钠缓冲液1：9倍稀释，搅拌15min，4℃静置10小时，抗体回收率为90％，去脂率为99％；该提取液用40％饱和度的硫酸铵盐析，得到的盐析液抗体回收率为82％，纯度达到69％。盐析液经截留分子量为100KD的有机陶瓷膜超滤后，截留液抗体回收率达到92％。纯度为85％。     

Cross-reactivity of anti-chicken IgY antibody with immunoglobulins of exotic avian species

Background: A major challenge in the serologic diagnosis of infectious diseases in exotic birds is the limited availability of species-specific antibodies. Objectives: The purpose of the current study was to determine if there is cross reactivity between commercially available anti-chicken IgY antibodies and immunoglobulins of several avian species, with particular emphasis on psittacines. Methods: To quantitate the reactivity with anti-chicken IgY, Western blot analysis was performed using plasma samples from many different avian species. Results were compared with gamma globulin fraction quantitation obtained by protein electrophoresis. Results: By Western blot, 2 protein bands corresponding to the heavy and light chains of chicken IgY were identified in species from 21 avian orders using 1 of 2 rabbit anti-chicken IgY antibodies. Densitometric analysis showed that the amount of immunoglobulin estimated from Western blots correlated strongly with data from protein electrophoresis assays. Conclusions: The results demonstrate that some commercially available anti-chicken IgY antibodies exhibit good cross-reactivity with most avian species.     

禽类免疫球蛋白

免疫球蛋白是存在于人和动物血清、组织液及其他外分泌液中的一类具有相似结构的球蛋白,它是抗体的本质。目前禽类中主要存在IgY、IgM、IgA三种免疫球蛋白,功能各不相同,在免疫系统中扮演着重要的角色。本文就禽类免疫球蛋白的结构与功能、生物效应和产生规律方面进行简要介绍。     

Protection of Fenneropenaeus chinensis (Osbeck, 1765) against the white spot syndrome virus using specific chicken egg yolk immunoglobulins by oral delivery

Two kinds of specific chicken egg yolk immunoglobulins (IgYs), IgY‐WSSV and IgY‐VP28, were, respectively, raised against the 2 mM binary ethylenimine (BEI)‐inactivated white spot syndrome virus (WSSV) and a principal envelope protein VP28. The activity of purified specific IgYs was stable under the conditions of 20–70 °C, pH 3.0–10.0 and 0–700 g L^?1 sucrose solution. In the neutralization assay, these high‐affinity IgY antibodies can specifically bind with the virus particles to protect shrimp (Fenneropenaeus chinensis) against WSSV infection. After oral delivery for 20 days, the IgY‐WSSV exerted a higher protection effect (RPS: 71.5%) than IgY‐VP28 (RPS: 63.7%). Moreover, an increase in RPS (79.2%) was found on addition of IgY‐WSSV:VP28 (0.1% IgY‐VP28 plus 0.2% IgY‐WSSV). This may indicate that neutralization of WSSV refers to the multiple‐hit model. By time‐course study of the levels of the specific IgYs in vivo, the data showed that the titre was enhanced to a relatively high level (P/N=8.35±0.45) at 3 days post administration, declined slightly (P/N=7.13±1.01) at 7 days post administration and then remained stable for further investigation. The stable antibody level potentially contributes towards blocking a large number of WSSV particles from entering and infecting on the major tissues at the early and late stages after challenge in shrimp.     

N-羟乙酰神经氨酸IgY抗体的制备及鉴定

本试验旨在建立N-羟乙酰神经氨酸(Neu5Gc) IgY抗体的制备及鉴定方法。通过碳二亚胺法将Neu5Gc和载体蛋白进行偶联,制备Neu5Gc人工完全抗原,用制备的完全抗原免疫海兰褐蛋鸡制备Neu5Gc-IgY抗体,经ELISA检测分析鉴定抗体活性。经紫外光谱扫描、琼脂糖凝胶电泳法初步判断Neu5Gc人工完全抗原制备成功,获得的特异性Neu5Gc-IgY抗体经ELISA检测分析,首次免疫后第7天产生抗体,抗体效价呈动态变化,随着免疫次数的增多,血清效价和抗体效价逐步上升,至初次免疫后63 d达到高峰,效价达1:10 000,停止加强免疫后抗体效价可持续一个月左右,1号鸡产生的IgY抗体具有很好的抗原竞争活性,获得竞争回归方程y=30.28x+16.923,线性系数R²=0.9581。以上结果表明,本试验制备的Neu5Gc IgY抗体取得了理想效果,为红肉、牛奶及肿瘤组织中Neu5Gc的检测奠定了基础。