鸡卵黄乙肝病毒特异性转移因子的制备及鉴定

目的制备鸡卵黄乙肝病毒特异性转移因子(EYHBV-TF),并检测其免疫活性。方法用乙肝疫苗免疫母鸡,收集鸡蛋,取卵黄进行透析,制备EYHBV-TF,参照药典要求,采用光谱法、高效液相色谱法(HPLC)、Lowry法等检测其理化性质。采用白细胞粘附抑制法(LAI)和迟发型皮肤超敏试验(DTH)检测特异免疫活性。结果EYHBV-TF是一种可透析、含有18种氨基酸、相对分子质量小于12000的小分子物质,其多肽含量为1·2mg/ml,核糖含量为152·94μg/ml,pH6·9±0·5,蛋白反应阴性,最大紫外吸收光谱在270nm处。该EYHBV-TF能明显抑制小鼠白细胞粘附(P<0·01),并能诱导小鼠跖趾部皮肤的迟发型超敏反应(P<0·01),与猪脾淋巴细胞制取的乙肝特异性转移因子(PSHBV-TF)检测结果接近(P>0·05)。而正常卵黄提取液组、非特异性转移因子和甲肝抗原(HAAg)对照组均不能表现出这两种效应(P>0·05)。结论EYHBV-TF与PS-TF主要理化性质和作用相类似,均具有乙肝抗原特异性细胞免疫活性。

Preparation and Characterization of Egg Yolk-derived Hepatitis B Virus Specific Transfer Factor

Objective To prepare egg yolk-derived hepatitis B virus specific transfer factor(EY_(HBV)-TF) and determine its immunological activity.Methods Immunize hens with hepatitis B vaccine and collect the eggs.Extract egg yolk and dialyze to prepare EY_(HBV)-TF,then study its physical and chemical properties by spectral analysis,HPLC and Lowry method et al.according to the requirements in Chinese Pharmacopoeia.Determine the immunological activity of EY_(HBV)-TF by leukocyte adherence inhibition(LAI) and delayed type hypersensitivity tests.Results EY_(HBV) -TF was a dialyzable substance(pH 6.9±0.5) with a relative molecular weight of less than 12 000,containing 18 kinds of amino acids.The polypeptide and ribose contents of EY_(HBV-TF were 1.2 mg/ml and 152.94 μg/ml respectively .The EY_(HBV)-TF was negative in protein reaction test and showed the maximum ultraviolet absorption at 270 nm.It inhibited the leukocyte adherence in mice significantly(P<0.01) and induced the DTH in metatarsophalangeal skin of mice (P<0.01) .The LAI and DTH effects of EY_(HBV)-TF showed no significant difference from those of pig splenic lymphocyte-derived hepatitis B virus specific transfer factor(PS_(HBV)-TF,P>0.05).However,the extract of normal egg yolk,non-specific transfer factor and hepatitis A antigen showed no LAI or DTH effects(P>0.05).Conclusion The major physical and chemical property and effect of EY_(HBV)-TF were similar to those of PS_(HBV)-TF.Both of them showed hepatitis B specific cell immunological activity.