Anderson George P, Liu Jinny L, Hale Martha L, Bernstein Rachael D, Moore Martin, Swain Marla D, Goldman Ellen R
Center for Bio/Molecular Science and Engineering, Naval Research Laboratory, 4555 Overlook Avenue SW, Washington, DC 20375, USA.
Anal Chem. 2008 Dec 15;80(24):9604-11. doi: 10.1021/ac8019398.
Single domain antibodies (sdAb) that bind ricin with high affinity and specificity were selected from a phage display library derived from the mRNA of heavy chain antibodies obtained from lymphocytes of immunized llamas. The sdAb were found to recognize three distinct epitopes on ricin. Representative sdAb were demonstrated to function as both capture and tracer elements in fluid array immunoassays, a limit of detection of 1.6 ng/mL was obtained. One sdAb pair in particular was found to be highly specific for ricin. While polyclonal antibodies cross react strongly with RCA120, the sdAb pair had minimal cross reactivity. In addition, the binders were found to be thermal stable, regaining their ricin binding activity following heating to 85 degrees C for an hour. Cycles of thermally induced unfolding of the sdAb and their subsequent refolding upon cooling was monitored by circular dichroism. As several of the sdAb were observed to bind to ricin's A chain, cell free translation assays were performed to monitor the ability of the sdAbs to inhibit ricin's biological activity. One of the sdAb (C8) was particularly effective and blocked ricin's biological activity with an effectiveness equal to that of a mouse antiricin antibody. These results indicate that antiricin sdAb have great potential for both diagnostic and therapeutic applications.
从免疫羊驼淋巴细胞中获得的重链抗体mRNA构建的噬菌体展示文库中筛选出了能与蓖麻毒素高亲和力和特异性结合的单域抗体（sdAb）。发现这些sdAb能识别蓖麻毒素上三个不同的表位。已证明代表性的sdAb在液相芯片免疫分析中可同时作为捕获和示踪元件，检测限达1.6 ng/mL。尤其发现一对sdAb对蓖麻毒素具有高度特异性。虽然多克隆抗体与RCA120有强烈的交叉反应，但这对sdAb的交叉反应极小。此外，还发现这些结合物具有热稳定性，在85℃加热1小时后仍能恢复其蓖麻毒素结合活性。通过圆二色性监测sdAb的热诱导解折叠循环及其冷却后的再折叠情况。由于观察到几种sdAb与蓖麻毒素的A链结合，因此进行了无细胞翻译试验以监测sdAb抑制蓖麻毒素生物活性的能力。其中一种sdAb（C8）特别有效，其阻断蓖麻毒素生物活性的效果与小鼠抗蓖麻毒素抗体相当。这些结果表明，抗蓖麻毒素sdAb在诊断和治疗应用方面具有巨大潜力。
