In situ hybridization versus northern analysis: working towards the correlation of two quantitative techniques for opioid and vasopressin mRNAs in the rat hypothalamus and pituitary.

Several physical and technical aspects of in situ hybridization are examined to help ascertain to what extent the total specific mRNA population of interest in a tissue section is actually detected using complementary RNA probes. Two experimental paradigms, Rot curve hybridization analysis and varying the tissue section thickness, indicate that complementary RNA (cRNA) probe hybridization to a specific mRNA in a tissue section can be made saturable, but remains so only for those mRNAs within a limited distance from the surface of the section. Direct comparison with Northern analysis, however, shows that in situ hybridization still gives comparable relative quantitative results for mRNA levels changing due to physiological stimulation.