Specificity of riboprobes for intracellular RNA in hybridization histochemistry

The specificity of the probe for intracellular RNA during hybridization histochemistry is usually confirmed by loss of the signal following ribonuclease pre-treatment of the target. Interpretation of such controls is complicated when single-stranded (asymmetric) RNA probes (riboprobes) are used for in situ hybridization, since the probes themselves may be degraded by residual ribonuclease. A protocol using the Ca2+-dependent enzyme, micrococcal nuclease, is presented to circumvent these difficulties.