QOPNA, Mass spectrometry center, Department of Chemistry, University of Aveiro, Portugal.
J Proteomics. 2012 Sep 18;75(17):5140-65. doi: 10.1016/j.jprot.2012.05.045. Epub 2012 Jul 15.
The present study aimed the evaluation of saliva sample pre-treatment, in particular the sample clearance usually performed by centrifugation, to the contribution of salivary proteome and peptidome. Using in-gel and off-gel approaches, a large content of salivary proteins was detected in the pellet fraction that is usually discarded. In addition, chaotropic/detergent treatment in combination with sonication, before the centrifugation step, resulted in salivary complex disruption and consequently in the extraction of high amounts of proteins. Based on this data, we suggest the use of urea/detergent with sonication as a standard saliva sample pre-treatment procedure. We also described a procedure to extract salivary peptides which can be performed even after saliva sample treatment with chaotropic/detergents. In overall, we reported for the first time the contribution of the pellet fraction to the whole saliva proteome. iTRAQ analysis highlighted a higher number of different peptides as well as distinct quantities of each protein class when after sample treatment with urea and sonication, acetone precipitation followed by solubilization with acetonitrile/HCl was performed.
本研究旨在评估唾液样本预处理，特别是通常通过离心进行的样本清除，对唾液蛋白质组和肽组的贡献。使用胶内和胶外方法，在通常丢弃的沉淀部分检测到大量唾液蛋白质。此外，在离心步骤之前，使用离液剂/去污剂处理并结合超声处理，导致唾液复合物破裂，从而提取大量蛋白质。基于这些数据，我们建议使用尿素/去污剂加超声处理作为唾液样本预处理的标准程序。我们还描述了一种提取唾液肽的方法，即使在使用离液剂/去污剂处理唾液样本后，也可以进行这种方法。总的来说，我们首次报道了沉淀部分对全唾液蛋白质组的贡献。iTRAQ 分析表明，当使用尿素和超声处理样品后，采用丙酮沉淀，然后用乙腈/盐酸溶解时，与使用离液剂/去污剂处理相比，不同的肽以及每种蛋白质类别的数量更多。
