Synthesis and characterisation of immunogens for the production of antibodies against small hydrophobic molecules with biosignature properties.
Synthesis and characterisation of immunogens for the production of antibodies against small hydrophobic molecules with biosignature properties.
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免疫原的合成和表征，用于生产针对具有生物特征的疏水性小分子的抗体。
DOI：
10.1016/j.aca.2011.09.021
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发表时间：
2011
影响因子：
6.2
通讯作者：
Sathe M
中科院分区：
文献类型：
--
作者：
Sathe M
在目前的研究中，五种不同类型的小疏水分子靶标，非典型抗体的产生，被结构修饰，以引入合适的反应功能和/或间隔，允许共价偶联到载体蛋白，从而产生稳定的载体-半抗原复合物。在生命标记芯片（LMC）的开发背景下，这些目标被选中作为现存和/或灭绝生命的标记。LMC是一种基于抗体的仪器，由英国领导的国际财团开发，将搭载ESA/NASA火星探测ExoMars联合任务飞往火星。半抗原蛋白偶联物被设计成在LMC发展的后续阶段用作抗体产生的免疫原和免疫测定试剂。由半抗原共价附着引起的蛋白质修饰程度由两种独立的方法确定，即三硝基苯磺酸（TNBSA）滴定剩余的蛋白质活性基团和基质辅助激光解吸/电离质谱（MALDI-MS）测定所得半抗原-蛋白质偶联物。在进一步的质量验证步骤中，将偶联物提交给动物免疫系统，获得具有中等特异性的多克隆抗体滴度。这些结果表明，本文所述的缀合物可以成功地用于产生针对小疏水分子的抗体。
In the present study, five different classes of small hydrophobic molecular targets, atypical for antibody generation, were structurally modified in order to introduce suitable reactive functionalities and/or spacers which allow covalent coupling to a carrier protein resulting in a stable carrier–hapten complex. These targets were chosen to serve as markers of extant and/or extinct life in the context of the development of the Life Marker Chip (LMC), an antibody-based instrument, which is being developed by a UK-led international consortium for flight to Mars on board the joint ESA/NASA Mars exploration ExoMars mission. The hapten–protein conjugates were designed to be used as immunogens for antibody generation and immunoassay reagents in subsequent stages of the LMC development. The extent of protein modification due to covalent attachment of hapten was determined by two independent methods, i.e. trinitrobenzenesulfonic acid (TNBSA) titrations of remaining protein reactive groups and matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) of the resultant hapten–protein conjugates. In a further quality validation step, the conjugates were presented to an animal's immune system and polyclonal antibody titres with moderate specificity were obtained. These results suggest that conjugates synthesized as described herein can successfully be used in the generation of antibodies targeting small hydrophobic molecules.
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