Table 15.6 Fluorescent detection reagents for imaging mitochondria and lysosomes.

                           Organic Dyes
(e.g., MitoTracker and LysoTracker dyes)
BacMam-Based Fluorescent Proteins
(e.g., CellLight reagents)
How they workPositively charged MitoTracker dyes localize to actively respiring mitochondria; weakly basic LysoTracker dyes accumulate in compartments with low pH.Combine targeting sequence–fluorescent protein fusion with the transduction efficiency of BacMam to label organelles independently of function (i.e., pH, mitochondrial membrane potential).Recognize specific target of interest (e.g., LAMP1, a lysosomal protein).
ApplicationsLive-cell imaging applications; fixable and thus compatible with antibody-based imaging applications.*Live-cell imaging applications; fixable and thus compatible with antibody-based imaging applications.*†Imaging fixed cells or tissue. Compatible with labeling with MitoTracker, LysoTracker, and CellLight reagents.
Typical workflowIncubate cells with the MitoTracker or LysoTracker reagent for approximately 5–30 min.The ready-to-use CellLight reagent is added to live cells, followed by an overnight incubation to allow for protein expression.Cells are fixed and permeabilized, then incubated with the antibody for labeling, and visualized with a fluorescently labeled secondary antibody.‡
* Please consult the product manual or contact technical service for additional information on the fixability of these reagents. † With fluorescent protein constructs, anti-GFP or anti-RFP antibodies can be used to amplify fluorescence signals. ‡ Secondary antibody is required if the primary antibody is not directly labeled.