Blocking and detection chemistries affect antibody performance on reverse phase protein arrays
Blocking and detection chemistries affect antibody performance on reverse phase protein arrays
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DOI：
10.1002/pmic.200700676
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发表时间：
2008-06-01
期刊：
影响因子：
3.4
通讯作者：
Olive, D. Michael
中科院分区：
文献类型：
--
作者：
Ambroz, Kristi L. H.;Zhang, Yonghong;Olive, D. Michael
抗体特异性是RP蛋白阵列（RPA）的关键。在Western blots和RPA中评估阻断和检测化学对抗体特异性的影响。阻断缓冲液显著影响Western blots上的非特异性条带，对阵列也有相应的影响。Tyramide信号放大（TSA）增加了western和阵列上的特异性和非特异性信号，掩盖了信号强度的预期梯度。这些结果表明，应该采用一致的阻断和检测条件进行抗体验证和后续的RPA实验。
Antibody specificity is critical for RP protein arrays (RPA). The effects of blocking and detection chemistries on antibody specificity were evaluated for Western blots and RPA. Blocking buffers significantly affected nonspecific banding on Western blots, with corresponding effects on arrays. Tyramide signal amplification (TSA) increased both specific and nonspecific signals on Westerns and arrays, masking the expected gradations in signal intensity. These results suggest that consistent blocking and detection conditions should be used for antibody validation and subsequent RPA experiments.