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首页> 外文期刊>Biotechnology Progress >Factors important in the extraction,stability and in vitro assembly of the hepatitis B surface antigen derived from recombinant plant systems
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Factors important in the extraction,stability and in vitro assembly of the hepatitis B surface antigen derived from recombinant plant systems

机译：来源于重组植物系统的乙型肝炎表面抗原的提取，稳定性和体外组装的重要因素

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The expression of vaccine antigens in edible plant material together with their delivery by the oralroute constitutes a powerful paradigm,with the potential to dramatically reduce the cost of vaccine production and administration,in addition to imporving distribution and patient compliance.These products will be subject to many of the same regualtions applied to current injectable vaccines,soreliable methods to quantify antigen and ensure stability in crude plant extracts are required.As a model system the hepatitis B surface antigen(HBsAg) was expressed in soybean and tobacco cell cultures.This complex antigen consists of membrane-associated small surface antigen proteins (p24~s),disulfide cross-linked to yield dimers and higher multimers.Although the total p24~s extracted from plant cells was relatively unaffected by detergent concentration,the quantification of antigenically reactive product depended strongly on the ratio of detergent to cell concentration.Furthermore,1-20% w/v sodium ascorbate imporved the measured levels of monoclonal-reactive antigen 4- to 12-fold.Detergent also influenced antigenstability in cell lysates stored at 4degC;under optimum condered the antigen susceptible to proteolytic degradation.This proteolysis could be counteracted by the addition of skim milk or its protein component,which stabilized antigenically reactive p24~s for up to 2 months.The imunologically relevant epitopes of HBsAg are critically dependent on disulfide bonding.By altering the sodium ascorbate concentration or buffer pH the proportion of HBsAg displaying the monoclonal reactive epitopes was increased between 8-and 20-fold.In addition,under certain conditions the dimerized p24~scould be convertd to oligomeric aggregates,resembling the form of the serum-derived antigen.These simple in vitro manipulations,compatible with the goal of a minimally processed oralvaccine,may prove valuable in increasing the immunogenicity of the plant-derived antigen.
机译：疫苗抗原在食用植物材料中的表达以及通过口服途径的传递构成了强大的范例，除了可以改善分布和患者依从性外，还可以大大降低疫苗生产和管理的成本。目前注射用疫苗有许多相同的规定，需要有选择的方法定量抗原并确保植物粗提物中的稳定性。作为模型系统，乙型肝炎表面抗原（HBsAg）在大豆和烟草细胞培养物中表达。这种复杂的抗原由膜相关的小表面抗原蛋白（p24〜s），二硫键交联以产生二聚体和更高的多聚体组成。虽然从植物细胞中提取的总p24〜s相对不受去污剂浓度的影响，但抗原反应性产物的定量取决于强烈地影响去污剂与细胞浓度的比率。此外，1-20％w / v sodi um抗坏血酸可将单克隆反应性抗原的测定水平提高4到12倍。洗涤剂还影响4degC储存的细胞裂解物中的抗原稳定性;在最佳条件下，抗原可降解为易于蛋白水解的蛋白，可通过添加脱脂奶来抵消这种蛋白水解作用或它的蛋白质成分，可以使抗原反应性p24〜s稳定长达2个月。HBsAg的免疫学相关表位严重依赖于二硫键。通过改变抗坏血酸钠的浓度或缓冲液pH，显示单克隆反应性表位的HBsAg比例为增加了8到20倍。此外，在某些条件下，二聚化的p24〜应转化为寡聚体，类似于血清来源抗原的形式。这些简单的体外操作，与最低限度加工的目标兼容口服疫苗可证明对提高植物来源抗原的免疫原性具有重要价值。

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