Towards a human proteome atlas:: High-throughput generation of mono-specific antibodies for tissue profiling
Towards a human proteome atlas:: High-throughput generation of mono-specific antibodies for tissue profiling
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DOI：
10.1002/pmic.200500072
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发表时间：
2005-11-01
期刊：
影响因子：
3.4
通讯作者：
Uhlen, M
中科院分区：
文献类型：
--
作者：
Nilsson, P;Paavilainen, L;Uhlen, M
系统地产生特异性抗体以探索人类蛋白质组存在巨大的需求。在这里，我们表明，可以产生的抗体特异性的人蛋白质的高通量的方式，涉及严格的亲和纯化使用重组蛋白表位签名标签（PrEST）作为免疫原和亲和配体。所产生的亲和试剂，这里称为单特异性抗体（msAb）的特异性，验证了一种新的蛋白质微阵列测定。通过蛋白质阵列测定判断，针对人蛋白质产生的464种抗体的成功率超过90%。使用从48个人组织产生的组织微阵列，将抗体用于患者活检的平行分析。与充分表征的单克隆抗体的比较分析显示相同或相似的特异性和表达模式。结果表明，一个全面的图集包含广泛的蛋白质表达和亚细胞定位数据的人类蛋白质组可以产生一个有效的方式与单特异性抗体。
A great need exists for the systematic generation of specific antibodies to explore the human proteome. Here, we show that antibodies specific to human proteins can be generated in a high-throughput manner involving stringent affinity purification using recombinant protein epitope signature tags (PrESTs) as immunogens and affinity-ligands. The specificity of the generated affinity reagents, here called mono-specific antibodies (msAb), were validated with a novel protein microarray assay. The success rate for 464 antibodies generated towards human proteins was more than 90% as judged by the protein array assay. The antibodies were used for parallel profiling of patient biopsies using tissue microarrays generated from 48 human tissues. Comparative analysis with well-characterized monoclonal antibodies showed identical or similar specificity and expression patterns. The results suggest that a comprehensive atlas containing extensive protein expression and subcellular localization data of the human proteome can be generated in an efficient manner with mono-specific antibodies.