Broihier Heather
Cold Spring Harb Protoc. 2012 Aug 1;2012(8):pdb.prot069542. doi: 10.1101/pdb.prot069542.
Whole-mount RNA in situ hybridizations with digoxigenin-conjugated probes and alkaline phosphatase biochemistry have been used widely for many years to map expression pattern domains in the Drosophila embryo. To capitalize on the number of molecular markers in the central nervous system (CNS) and to enable expression analysis at the single-cell level, fluorescence in situ hybridization procedures are becoming standard. This protocol describes methods for the simultaneous detection of RNA and protein using fluorescence in Drosophila embryos. It uses the tyramide signal amplication (TSA) system from PerkinElmer to amplify a horseradish peroxidase (HRP) signal. By combining this technology with an HRP-conjugated antidigoxigenin antibody, we can detect standard antidigoxigenin RNA probes fluorescently.
多年来，地高辛配体偶联探针的全胚胎RNA原位杂交和碱性磷酸酶生化方法已被广泛用于绘制果蝇胚胎中的表达模式域。为了利用中枢神经系统（CNS）中的分子标记数量并实现单细胞水平的表达分析，荧光原位杂交程序正成为标准方法。本方案描述了在果蝇胚胎中使用荧光同时检测RNA和蛋白质的方法。它使用珀金埃尔默公司的酪胺信号放大（TSA）系统来放大辣根过氧化物酶（HRP）信号。通过将该技术与HRP偶联的抗地高辛抗体相结合，我们可以荧光检测标准的抗地高辛RNA探针。
