Scenario description
For this scenario the assay simultaneously detects the expression pattern of two genes of interest. In this scenario both the expression level and the percent of cells expressing each of the targets could be of importance and therefore can be analyzed according to the singleplex scenarios applicable for each of the targets (Figures 12 and 13).
Figure 12. Drd1 (red) and Drd2 (green) expression on distinct medium spiny neuronal cell populations in the striatum of normal mouse brain using the RNAscope ® Multiplex Fluorescent Assay. Cells are counterstained with DAPI (blue).
Figure 13. Cnr1 (green) and Drd1 (red) expression in the hippocampus of normal mouse brain using the RNAscope ® Multiplex Fluorescent Assay. Cells are counterstained with DAPI (blue).
Analysis guidelines
To determine the target expression for each of the genes within a particular cell type, one can qualitatively and quantitatively assess the expression pattern by using methodologies #1, #2 and #3. These methodologies can be combined to create percentage of cells positive. Cells can be scored visually at 40x based on number of cells with >1 dot/cell across the entire tissue section. Percent dual positive can be calculated in two ways (Table 4):
