In Situ Polymerase Chain Reaction and Hybridization to Detect Low‐Abundance Nucleic Acid Targets

This unit presents a novel approach for detecting low-abundance nucleic acid targets in nuclear and cytoplasmic regions by amplification of specific target sequences using an in situ polymerase chain reaction (ISPCR). If the target sequence is RNA, ISPCR is preceded by in situ reverse transcription. Following ISPCR, in situ hybridization is performed. An Alternate Protocol describes a variant in situ method for simultaneously reverse transcribing and amplifying RNA transcripts using recombinant Thermos thermophilos (rTth) polymerase. In situ hybridization and detection of amplified targets is also described.