Science for Life Laboratory, KTH - Royal Institute of Technology, SE-171 21, Stockholm, Sweden.
Department of Protein Science, KTH - Royal Institute of Technology, SE-106 91, Stockholm, Sweden.
Nat Commun. 2018 Oct 8;9(1):4130. doi: 10.1038/s41467-018-06642-y.
There is a need for standardized validation methods for antibody specificity and selectivity. Recently, five alternative validation pillars were proposed to explore the specificity of research antibodies using methods with no need for prior knowledge about the protein target. Here, we show that these principles can be used in a streamlined manner for enhanced validation of research antibodies in Western blot applications. More than 6,000 antibodies were validated with at least one of these strategies involving orthogonal methods, genetic knockdown, recombinant expression, independent antibodies, and capture mass spectrometry analysis. The results show a path forward for efforts to validate antibodies in an application-specific manner suitable for both providers and users.
需要标准化的抗体特异性和选择性验证方法。最近，提出了五种替代验证支柱，使用无需事先了解蛋白质靶标的方法来探索研究抗体的特异性。在这里，我们表明，这些原则可以以简化的方式用于增强 Western blot 应用中研究抗体的验证。使用涉及正交方法、基因敲低、重组表达、独立抗体和捕获质谱分析的这些策略中的至少一种，对 6000 多种抗体进行了验证。结果表明，朝着以适合提供者和用户的特定于应用的方式验证抗体的努力迈进了一步。
