Qian X, Bauer R A, Xu H S, Lloyd R V
Department of Laboratory Medicine and Pathology, Mayo Clinic and Mayo Foundation, Rochester, Minnesota, USA.
Appl Immunohistochem Mol Morphol. 2001 Mar;9(1):61-9.
In situ hybridization (ISH) is a powerful molecular tool used to visualize nucleic acids in tissues and cells. However, its use is limited by the relative lack of sensitivity in detecting low copy numbers of nucleic acids. Several strategies have been developed to improve the threshold levels of in situ detection of nucleic acid by amplification of either target nucleic acid sequences before ISH (such as in situ polymerase chain reaction) or after the hybridization procedure (such as tyramide signal amplification [TSA]). The authors compared the use of different types of probes to detect calcitonin mRNA in 10 cases of formalin-fixed, paraffin-embedded medullary thyroid carcinoma with and without TSA. In addition, dot blot hybridization was used to compare the signal amplification by TSA with oligonucleotide. cDNA, and cRNA probes. These results show that cRNA probes are the most sensitive types of probes for detecting mRNA molecules in formalin-fixed, paraffin-embedded tissue sections and that tyramide amplification can increase the sensitivity for detection of calcitonin mRNA in formalin-fixed, paraffin-embedded tissue sections at least 2- to 4-fold with cRNA probes.
原位杂交（ISH）是一种用于在组织和细胞中可视化核酸的强大分子工具。然而，其应用受到检测低拷贝数核酸时相对缺乏敏感性的限制。已经开发了几种策略来通过在ISH之前扩增靶核酸序列（如原位聚合酶链反应）或在杂交程序之后（如酪胺信号放大[TSA]）来提高核酸原位检测的阈值水平。作者比较了在10例福尔马林固定、石蜡包埋的甲状腺髓样癌中使用不同类型探针并结合或不结合TSA检测降钙素mRNA的情况。此外，使用斑点印迹杂交来比较TSA与寡核苷酸、cDNA和cRNA探针的信号放大情况。这些结果表明，cRNA探针是在福尔马林固定、石蜡包埋的组织切片中检测mRNA分子最敏感的探针类型，并且酪胺放大可以使cRNA探针在福尔马林固定、石蜡包埋的组织切片中检测降钙素mRNA的敏感性至少提高2至4倍。
