IMAGING TO MEASURE LOCALIZATION OF PROTEIN IN THE NUCLEAR COMPARTMENT
In the simple schematic shown to the right, untreated cells express a moderate amount of the marker protein (depicted in green), which appears to be cytoplasmic. The nuclei (depicted in blue) are typically labelled using Hoechst 33342 (learn why we use Hoechst here).
In this schematic, one condition leads to an increase in cytoplasmic levels while the other leads to accumulation of the protein marker in the nucleus (colocalization of the blue and green signals is depicted in cyan).
Because we use the Hoechst marker to demark the location of the nucleus, we are able to measure the amount of the protein marker that is coincident with it.
Using this approach, it is possible to quantify changes in protein expression level AND cytoplasmic-nuclear localization for 4-5 markers in a multiplexed fashion relative to the nuclear signal.