Evaluation of ethanol-fixed, paraffin-embedded tissues for proteomic applications
Evaluation of ethanol-fixed, paraffin-embedded tissues for proteomic applications
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DOI：
10.1002/pmic.200390056
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发表时间：
2003-04-01
期刊：
影响因子：
3.4
通讯作者：
Emmert-Buck, MR
中科院分区：
文献类型：
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作者：
Ahram, M;Flaig, MJ;Emmert-Buck, MR
我们先前报道，乙醇固定和石蜡包埋的组织可以产生良好的组织形态和良好的大分子保存。在这里，我们对乙醇固定的组织进行了详细的蛋白质组学评价。从乙醇固定、石蜡包埋的前列腺组织中提取蛋白质，经双向凝胶电泳法(2-DE)分离，并用标准方法染色后，可成功地检测到数百个蛋白质分子并成功地进行了质谱分析。从乙醇固定的组织中获得的蛋白质图谱与从冰冻组织中观察到的高度相似，而从福尔马林固定的材料中获得的蛋白质回收很差。用激光捕获显微切割从乙醇固定的组织切片中分离的特定细胞的蛋白质含量也可以用2-DE成功地分析。我们观察到，组织切片的伊红染色对蛋白质分离有不利影响，而苏木精染色的影响很小。为了说明乙醇固定的组织在蛋白质组发现研究中的适用性，我们比较了患者匹配的正常前列腺上皮细胞和从乙醇固定的石蜡包埋组织中获得的侵袭性腺癌细胞的蛋白质谱。通过质谱学分析，发现并鉴定了一些差异表达的蛋白质。对乙醇固定的组织切片进行的免疫组织化学分析与蛋白质组发现的结果是一致的。根据这些结果，我们得出结论，乙醇固定的组织可以成功地用于蛋白质组分析。
We previously reported that ethanol fixation and paraffin embedding of tissues produce excellent histomorphology and good preservation of macromolecules. Here, we present a detailed evaluation of ethanol-fixed tissues for proteomic initiatives. When proteins were extracted from ethanol-fixed, paraffin-embedded prostate tissue, resolved by two-dimensional gel electrophoresis (2-DE), and stained by standard methods, several hundred protein molecules could be detected and successfully analyzed by mass spectrometry. Protein profiles obtained from ethanol-fixed tissues were highly similar to those observed from frozen tissues, in contrast to the poor protein recovery from formalin-fixed material. The protein content of specific cells that were microdissected from ethanol-fixed tissue sections using laser capture microdissection could also be successfully analyzed by 2-DE. We observed that eosin staining of tissue sections had a detrimental effect on protein separation,,whereas hematoxylin staining had minimal consequence. In order to illustrate the applicability of ethanol-fixed tissues for proteomic discovery studies, we compared the protein profiles of patient-matched, normal prostatic epithelial cells and invasive adenocarcinoma cells obtained from ethanol-fixed, paraffin-embedded tissues. A number of differentially expressed proteins was discovered and identified by mass spectrometry. Immunohistochemical analyses performed on ethanol-fixed tissue sections were in agreement with the proteomic discovery findings. In light of these results, we conclude that ethanol-fixed tissues can be successfully utilized for proteomic analyses.