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) | Antibodies | |
|---|---|---|---|
| How they work | Positively 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). |
| Applications | Live-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 workflow | Incubate 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. | |||
For Research Use Only. Not for use in diagnostic procedures.