Generation of monoclonal antibodies to cryptic collagen sites by using subtractive immunization
Generation of monoclonal antibodies to cryptic collagen sites by using subtractive immunization
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DOI：
10.1089/02724570050198893
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发表时间：
2000-10-01
期刊：
影响因子：
--
通讯作者：
Brooks, PC
中科院分区：
文献类型：
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作者：
Xu, JS;Rodriguez, D;Brooks, PC
细胞外基质（ECM）在正常和病理过程的调节中发挥着重要作用。在ECM中发现的最丰富表达的组分是胶原蛋白。已知三螺旋胶原蛋白对除基质金属蛋白酶（MMP）家族成员以外的蛋白水解裂解具有高度抗性。迄今为止，关于胶原代谢对人类疾病的生物化学后果知之甚少。这部分是由于缺乏可以区分蛋白水解和三螺旋形式的胶原蛋白的特异性试剂。在这里，我们使用消减免疫（SI）技术，以产生两个独特的单克隆抗体（单克隆抗体HUIV26和HUI77），与变性和蛋白水解形式的胶原蛋白反应，但显示很少，如果有的话，与三螺旋胶原蛋白的反应。重要的是，HUIV26和HUI77与人黑色素瘤肿瘤ECM内的隐蔽位点反应，证明了它们用于体内免疫组织化学分析的实用性。因此，这些新的单克隆抗体的产生不仅确定三螺旋胶原蛋白内的特定隐蔽表位，而且还提供了重要的新试剂，用于研究正常以及病理过程中的胶原蛋白重塑的作用。
The extracellular matrix (ECM) plays a fundamental role in the regulation of normal and pathological processes. The most abundantly expressed component found in the ECM is collagen. Triple helical collagen is known to be highly resistant to proteolytic cleavage except by members of the matrix metalloproteinase (MMP) family of enzymes. To date little is known concerning the biochemical consequences of collagen metabolism on human diseases. This is due in part to the lack of specific reagents that can distinguish between proteolyzed and triple helical forms of collagen. Here we used the technique of Subtractive Immunization (SI) to generate two unique monoclonal antibodies (MAbs HUIV26 and HUI77) that react with denatured and proteolyzed forms of collagen, but show little if any reaction with triple helical collagen. Importantly, HUIV26 and HUI77 react with cryptic sites within the ECM of human melanoma tumors, demonstrating their utility for immunohistochemical analysis in vivo. Thus, the generation of these novel MAbs not only identify specific cryptic epitopes within triple helical collagen, but also provide important new reagents for studying the roles of collagen remodeling in normal as well as pathological processes.