抗痢疾志贺氏菌卵黄抗体的制备及其稳定性

目的制备特异性抗痢疾志贺氏菌的鸡卵黄免疫球蛋白,研究母鸡对痢疾志贺氏菌的免疫应答规律及抗体的稳定性。方法以两种不同剂量的痢疾志贺氏菌作为抗原免疫产卵母鸡,应用酶联免疫吸附测定法检测抗痢疾志贺氏菌IgY的效价及其稳定性。结果初次免疫后第12天,卵黄中开始出现抗痢疾志贺氏菌的IgY,免疫应答持续时间超过25周。稳定性试验表明IgY在pH3-11,温度不超过60℃,蔗糖质量浓度不高于700 g/L的渗透压条件下活性稳定。结论用痢疾志贺氏菌免疫母鸡制备的IgY效价高,理化性质稳定。     

抗须癣毛癣菌细胞壁蛋白的卵黄抗体制备和鉴定

目的：提取须癣毛癣菌的细胞壁蛋白作为免疫原制备特异性卵黄抗体,并鉴定其生物活性,为卵黄抗体在预防与治疗皮肤癣疾病的应用奠定基础。方法：本研究采用冷碱抽提的方法提取须癣毛癣菌的细胞壁蛋白,并用其免疫健康的产蛋母鸡。采用聚乙二醇两步沉淀法及饱和硫酸铵盐析提纯卵黄抗体;用Bradford法检测卵黄抗体中蛋白含量;SDS-PAGE凝胶电泳分析测定卵黄抗体的纯度以及相对分子质量;ELISA检测纯化的卵黄抗体的效价;Western blot分析特异性卵黄抗体的免疫反应性。结果：提取的卵黄抗体中蛋白纯度达到87.27%。由细胞壁蛋白制备的特异性卵黄抗体在初免20 d后效价开始升高,到45天达到最高值（1∶32 000）。Western blot结果显示由细胞壁蛋白制备的卵黄抗体能与其良好的特异性结合,具有较好的免疫反应性。结论：本实验提取的须癣毛癣菌细胞壁蛋白作为免疫原可以制备出特异性较强的卵黄抗体,为须癣毛癣菌感染的皮肤疾病提供了治疗新思路。     

抗肠道病毒71型特异性卵黄抗体的制备及鉴定

目的:制备特异性抗肠道病毒71型(Enterovirus 71,EV71)的卵黄抗体并检测其生物学性能,这对EV71感染手足口病的预防和治疗具有重大的意义。方法:用纯化并灭活后EV71作为抗原免疫健康产蛋鸡。采用本实验室水提综合聚乙二醇法提取纯化鸡卵黄抗体;用Bradford法测定卵黄抗体提取液的蛋白含量;还原性SDS-PAGE凝胶电泳分析测定提取蛋白的纯度及相对分子质量;分别用ELISA、双向免疫琼脂扩散检测纯化特异性卵黄抗体抗EV71效价;Western blot分析特异性卵黄抗体的免疫反应性。结果:卵黄中卵黄抗体含量达7.76 mg/ml,卵黄抗体的纯度为94.86%。初免10天后蛋黄中可检测到特异性抗EV71卵黄抗体,初免40天后ELISA检测抗体效价高达1∶20 480,双向琼脂扩散检测效价达1∶16。提取的卵黄抗体具有良好的免疫反应性,能与EV71特异性结合。结论:水提综合聚乙二醇法提取纯化得到的抗EV71卵黄抗体产量和纯度高、效价好且具有良好的免疫反应性。     

Growth inhibition of Staphylococcus aureus by chicken egg yolk antibodies

Introduction  

The aim was to evaluate the ability of egg yolk antibody (IgY) in blocking Staphylococcus aureus growth in vitro.     

抗人蔗糖酶卵黄抗体的制备及其稳定性和体外活性研究

目的:制备抗人蔗糖酶(Sucrase,SUC)卵黄抗体(egg yolk immunoglobulin Y,IgY)并对其稳定性、体外活性进行研究。方法:采用人SUC 蛋白为抗原,免疫海兰灰蛋鸡,水稀释和盐析法提取纯化IgY;间接ELISA 法监测抗体效价变化并评价其稳定性;SDS-PAGE 和Western blot 分析IgY 的纯度和特异性;PNPG 法测IgY 对琢鄄葡萄糖苷酶的体外抑制效果。结果:初次免疫10 d 后检测到IgY,40 d 后效价最高达到1 12 800;提取的IgY 重链和轻链分别在65 kD 和25 kD 处,且能够与抗原蛋白特异性结合;29 ~69益范围内水浴15 min 和pH 4 ~7 之间37益水浴4 h,抗体效价无变化;胰蛋白酶与胃蛋白酶作用IgY2 h,抗体效价降至一半,延长作用时间,IgY 对胃蛋白酶耐受力较胰蛋白酶耐受力更强;IgY 对琢鄄葡萄糖苷酶具有一定的抑制作用,呈现剂量相关性,半抑制浓度(IC50)值为0郾540 mg/ ml。结论:抗人蔗糖酶卵黄抗体(S-IgY)的成功制备为后续用于2型糖尿病大鼠模型体内实验研究奠定基础。     

Neutralization of enterotoxigenic escherichia coli heat‐labile toxin by chicken egg yolk immunoglobulin Y and its antigen‐binding fragments

The protective effect of chicken egg yolk immunoglobulin Y (IgY) and its antigen‐binding FAb’ fragments was studied by their neutralization of heat‐labile toxin (LT) produced by enterotoxigenic Escherichia coli (ETEC) strain H10407. High levels of specific antibodies against LT were produced and maintained over a period of 8 months in eggs of hyper‐immunized hens. The FAb’ fragment produced by peptic digestion was found to be as effective as the whole IgY molecule in neutralizing LT. Removal of the enterotoxin‐specific antibodies eliminated the neutralization activities of both IgY and the FAb’ fragment. Heat pasteurization at 65°C for 15 min and presence of high salt (1.5 M‐NaCl) at pH 7.0 did not change the neutralization activities of either IgY or FAb’. Although incubation at pH of 2.0 for 4 h led to a 16‐fold decrease in the neutralization activities of IgY and FAb’, substantial neutralization activities were retained. The combination of low pH and high salt content, however, resulted in the complete destruction of the neutralization activity of IgY, but not the FAb’ fragment.     

特异抗内毒素鸡蛋黄免疫球蛋白效价测定及理化性质研究

目的：研究特异性抗内毒素鸡蛋黄免疫球蛋白理化性质，探索一种防治肠源性感染，治疗：内毒素血症的新途径。方法：用大肠杆菌J5株(菌体表面类脂A裸露)作抗原免疫25周龄Roman鸡，制备特异性抗内毒素IgY，用酶联免疫吸附技术检测其效价及对热、酸、碱和酶的稳定性。结果：IgY抗体效价为1：25 600，能耐受巴氏消毒，对酸、碱、酶有很好的稳定性。结论：用大肠杆菌J5株免疫鸡制备的IgY效价高，理化性质稳定，适合口服应用。     

Influence of lysophosphatidylcholine,phosphatidylcholine, and hen egg yolk on contractile effects of acetylcholine on smooth muscles of rat stomach

Experiments on smooth muscles of rat stomach showed that lysophosphatidylcholine in concentrations of 10⁻⁸ and 10⁻⁷ g/ml does not modulate the tonotropic effect of acetylcholine (10⁻⁶ g/ml), in a concentration of 10⁻⁶ g/ml potentiated this effect (similarly to phosphatidylcholine, 10⁻⁶ g/ml), and reduced it in concentrations of 10⁻⁵−10⁻⁴ g/ml (similarly to hen egg yolk in dilutions of 1:500, 1:100, and 1:500). These data indicate that lysophosphatidylcholine modifies signal transduction from the receptor to G protein. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 143, No. 6, pp. 604–607, June, 2007     

The production of anti-Vibrio harveyi egg yolk immunoglobulin and evaluation of its stability and neutralisation efficacy

In order to replace the antibiotic treatment for control of Vibrio harveyi, a causal agent of luminous disease in Black tiger shrimp, anti-V. harveyi IgY was produced and showed its potential in our preliminary study. However, for further use as feed additive, the IgY stability should be evaluated. The titre of specific IgY was enhanced with an immunostimulant, C-phosphate guanosine oligodeoxynucleotide (CpG-ODN). IgY was stable at natural pH while the activity was decreased below pH 4 or above pH 10. The supplementation of 30% sorbitol significantly enhanced the IgY stability at tested temperatures in excess of 70°C. The activity of IgY remained at 9 and 94% after 4 h incubation in simulated gastric and intestinal fluids, respectively. A storage temperature at ?20°C was found to be the best condition. IgY at the concentrations of 1, 5 and 10 mg/ml efficiently inhibited V. harveyi in vitro within 24 h after exposure.