Cox Melissa L, Schray Carrie L, Luster Chandra N, Stewart Zachary S, Korytko Peter J, M Khan Kanwar Nasir, Paulauskis Joseph D, Dunstan Robert W
Pfizer Global Research and Development, Safety Sciences Ann Arbor, 2800 Plymouth Road, Tox 35/134, Ann Arbor, MI 48105, USA.
Exp Mol Pathol. 2006 Apr;80(2):183-91. doi: 10.1016/j.yexmp.2005.10.002. Epub 2005 Dec 5.
Molecular characterization of morphologic change requires exquisite tissue morphology and RNA preservation; however, traditional fixatives usually result in fragmented RNA. To optimize molecular analyses on fixed tissues, we assessed morphologic and RNA integrity in rat liver when sections were fixed in 70% neutral-buffered formalin, modified Davidson's II, 70% ethanol, UMFIX, modified Carnoy's, modified methacarn, Bouin's, phosphate-buffered saline, or 30% sucrose. Each sample was subjected to standard or microwave fixation and standard or microwave processing, and sections were evaluated microscopically. RNA was extracted and assessed for preservation of quality and quantity. Modified methacarn, 70% ethanol, and modified Carnoy's solution each resulted in tissue morphology representing a reasonable alternative to formalin. Modified methacarn and UMFIX best preserved RNA quality. Neither microwave fixation nor processing affected RNA integrity relative to standard methods, although morphology was modestly improved. We conclude that modified methacarn, 70% ethanol, and modified Carnoy's solution provided acceptable preservation of tissue morphology and RNA quality using both standard and microwave fixation and processing methods. Of these three fixatives, modified methacarn provided the best results and can be considered a fixative of choice where tissue morphology and RNA integrity are being assessed in the same specimens.
形态学变化的分子特征分析需要精细的组织形态学和RNA保存；然而，传统固定剂通常会导致RNA片段化。为了优化对固定组织的分子分析，我们评估了大鼠肝脏在分别用70%中性缓冲福尔马林、改良戴维森氏固定液II、70%乙醇、UMFIX、改良卡诺氏固定液、改良甲醇-冰醋酸固定液、布因氏固定液、磷酸盐缓冲盐水或30%蔗糖固定切片时的形态学和RNA完整性。每个样本都进行了标准或微波固定以及标准或微波处理，并通过显微镜对切片进行评估。提取RNA并评估其质量和数量的保存情况。改良甲醇-冰醋酸固定液、70%乙醇和改良卡诺氏固定液所得到的组织形态都代表了福尔马林的合理替代方案。改良甲醇-冰醋酸固定液和UMFIX能最好地保存RNA质量。相对于标准方法，微波固定和处理均未影响RNA完整性，不过形态学有适度改善。我们得出结论，使用标准和微波固定及处理方法时，改良甲醇-冰醋酸固定液、70%乙醇和改良卡诺氏固定液对组织形态和RNA质量的保存效果均可接受。在这三种固定剂中，改良甲醇-冰醋酸固定液效果最佳，在对同一标本同时评估组织形态和RNA完整性时可被视为首选固定剂。
