In situ hybridization to cellular RNA.

In situ hybridization to cellular RNA is used to determine the cellular localization of specific messages within complex cell populations and tissues. In this unit, protocols are described for hybridizing slide-mounted paraffin sections or cryosections with labeled probes. Support protocols describe synthesis of 35S-labeled riboprobes and dsDNA probes, which are then detected using film autoradiography or emulsion autoradiography. Another support protocol describes synthesis of digoxigenin-labeled RNA probes, which are non-radioactive and thus have several advantages. They are easily synthesized in large quantities, they are stable for several months, and they can be reused up to three times. An additional advantage of RNA versus DNA probes is that they result in cleaner signals because nonspecifically bound probe is removed during ribonuclease treatment.