Detection of biomolecules in tissues provides contextual information and the possibility to assess the interaction of different cell types and markers. Routine qualitative assessment of immune- and oligonucleotide-based methods in research and the clinic has been associated with assay variability because of lack of stringent validation and subjective interpretation of results. As a result, the vast majority of in situ assays in clinical usage are nonquantitative and, although useful, often of questionable scientific validity. Here, we revisit the reporters and methods used for single- and multiplexed in situ visualization of protein and RNA. Then we examine methods for the use of quantitative platforms for in situ measurement of protein and mRNA levels. Finally, we discuss the challenges of the transition of these methods to the clinic and their potential role as tools for development of companion diagnostic tests.