Tourtellotte W W, Verity A N, Schmid P, Martinez S, Shapshak P
J Virol Methods. 1987 Feb;15(2):87-99. doi: 10.1016/0166-0934(87)90052-8.
A novel method for covalently binding formalin fixed paraffin embedded (FFPE) tissue sections to glass microscope slides is validated suitable for in situ hybridization (ISH). Using the organosilane methodology of Maples (1985), 100% tissue adhesion is reported with no nonspecific probe binding, staining, or autoradiographic artefacts. JC viral nucleic acid sequences are successfully detected in FFPE progressive multifocal leukoencephalopathy brain tissue and the Tm of the hybridized product is estimated. From the Tm the most stringent washing condition resulting in an optimal signal to noise ratio is determined. A comparison is made between currently used methods of tissue adhesion and the proposed organosilane methodology. This methodology greatly facilitates studies of conditions for ISH and elucidation of mechanisms of viral infections requiring consecutive FFPE sections. It is also applicable to studies using cryosections and cultured cells.
一种将福尔马林固定石蜡包埋（FFPE）组织切片共价结合到玻璃显微镜载玻片上的新方法经验证适用于原位杂交（ISH）。采用Maples（1985年）的有机硅烷方法，报告称组织附着力达100%，且无非特异性探针结合、染色或放射自显影假象。在FFPE进行性多灶性白质脑病脑组织中成功检测到JC病毒核酸序列，并估算了杂交产物的熔解温度（Tm）。根据Tm确定了产生最佳信噪比的最严格洗涤条件。对目前使用的组织黏附方法与所提出的有机硅烷方法进行了比较。该方法极大地促进了ISH条件的研究以及对需要连续FFPE切片的病毒感染机制的阐明。它也适用于使用冷冻切片和培养细胞的研究。
