Use of reverse phase protein microarrays to study protein expression in leukemia: technical and methodological lessons learned.
Use of reverse phase protein microarrays to study protein expression in leukemia: technical and methodological lessons learned.
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DOI：
10.1007/978-1-61779-286-1_10
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发表时间：
2011
期刊：
影响因子：
--
通讯作者：
Coombes KR
中科院分区：
文献类型：
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作者：
Kornblau SM;Coombes KR
白血病非常适合通过RPPA进行蛋白质组学分析，这是由于来自血液或骨髓的原始细胞易于获得。在这篇综述中，我们回顾了影响RPPA数据质量的方法和程序问题。我们推荐接触式打印机，以最大限度地减少样品量和蒸发，并最大限度地增加每个载玻片的样品。还审查了样品选择和处理的影响。蛋白质最好在获取之日新鲜制备，因为冷冻保存会改变某些疾病中的蛋白质表达水平。还需要快速处理以避免磷酸化随时间的变化。只要使用白血病原始细胞富集程序，样本来源、血液与骨髓似乎不会影响结果。选择正确的“正常”对照对于比较患病与“正常”表达是重要的。各种手段规范化的数据进行了讨论。
Leukemias are well suited to proteomic profiling by RPPA due to the ready accessibility of blasts from the blood or marrow. In this review, we review methodological and procedural issues that affect the quality of RPPA data. We recommend contact printers that minimize sample quantities and evaporation and maximize sample per slide. The impact of sample selection and handling is reviewed as well. Protein is best prepared fresh on the date of acquisition as cryopreservation changes protein expression levels in some diseases. Rapid processing is also required to avoid changes in phosphorylation over time. Sample source, blood vs. marrow does not seem to affect results as long as leukemic blast enrichment procedures are utilized. The choice of the correct “normal” control is important for comparing diseased to “normal” expression. Various means of normalizing the data are discussed.