Signal Amplification for DNA and mRNA

In situ hybridization (ISH) permits the localization of specific unique or repeated DNA and RNA sequences at the level of individual cells (1 -4 ). It has significantly advanced the study of gene structure and expression, and, in addition to morphological identification of cell types involved, ISH also allows some quantification of observations, e.g., with respect to tumor burden or viral load. Despite its high degree of detection specificity, the technique still does not allow the routine detection of DNA sequences less than 5 kb in size, and in the case of tissue sections the detection sensitivity is even more limited. The threshold levels for mRNA detection are more difficult to determine, with the reported sensitivity limits of 1–20 copies of mRNA per cell being achieved only in the most sensitive protocols (5 ,6 ,6a ).