Evaluation of Tumor Interstitial Fluid-Extraction Methods for Proteome Analysis: Comparison of Biopsy Elution versus Centrifugation.,Journal of Proteome Research

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Evaluation of Tumor Interstitial Fluid-Extraction Methods for Proteome Analysis: Comparison of Biopsy Elution versus Centrifugation.
Journal of Proteome Research ( IF 3.8 ) Pub Date : 2019-12-26 , DOI: 10.1021/acs.jproteome.9b00770
Clara Matas-Nadal ₁ , Joan Josep Bech-Serra ₂ , Marta Guasch-Vallés _{1,

3} , Josep Manel Fernández-Armenteros _{1,

4} , Carla Barceló ₁ , Josep Manel Casanova _{1,

3,

4} , Carolina de la Torre Gómez ₂ , Rafael Aguayo Ortiz _{1,

4} , Eloi Garí _{1,

3}

Affiliation

The analysis of tumor interstitial fluid (TIF) composition is a valuable procedure to identify antimetastatic targets, and different laboratories have set up techniques for TIF isolation and proteomic analyses. However, those methods had never been compared in samples from the same tumor and patient. In this work, we compared the two most used methods, elution and centrifugation, in pieces of the same biopsy samples of cutaneous squamous cell carcinoma (cSCC). First, we established that high G-force (10 000g) was required to obtain TIF from cSCC by centrifugation. Second, we compared the centrifugation method with the elution method in pieces of three different cSCC tumors. We found that the mean protein intensities based in the number of peptide spectrum matches was significantly higher in the centrifuged samples than in the eluted samples. Regarding the robustness of the methods, we observed higher overlapping between both methods (77–80%) than among samples (50%). These results suggest that there exists an elevated consistence of TIF composition independently of the method used. However, we observed a 3-fold increase of extracellular proteins in nonoverlapped proteome obtained by centrifugation. We therefore conclude that centrifugation is the method of choice to study the proteome of TIF from cutaneous biopsies.

中文翻译：

用于蛋白质组分析的肿瘤间质液提取方法的评估：活检洗脱与离心比较。

肿瘤间质液（TIF）成分的分析是确定抗转移目标的有价值的程序，并且不同的实验室已经建立了TIF分离和蛋白质组学分析技术。但是，从未对来自相同肿瘤和患者的样品中的那些方法进行过比较。在这项工作中，我们在皮肤鳞状细胞癌（cSCC）的相同活检样本中比较了两种最常用的方法，即洗脱和离心。首先，我们确定了高G力（10000 g需要通过离心从cSCC获得TIF。其次，我们将离心方法与洗脱方法在三种不同的cSCC肿瘤中进行了比较。我们发现，基于肽谱匹配数的平均蛋白质强度在离心样品中显着高于洗脱样品。关于方法的鲁棒性，我们观察到两种方法之间的重叠率更高（77-80％），而样品之间的重叠率更高（50％）。这些结果表明，与所使用的方法无关，存在着提高的TIF组成的一致性。但是，我们观察到通过离心获得的不重叠蛋白质组中细胞外蛋白质增加了3倍。因此，我们得出结论，离心是研究皮肤活检中TIF蛋白质组的一种选择方法。

更新日期：2019-12-26

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