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Cell tracking dyes are non-toxic, stable probes that allow for cell movement and localization studies in live cells. CellTracker (short-term) and Qtracker (long-term) are passively loaded into cells and retained through several generations but not transferred to adjacent cells. These cell tracking dyes are available in a range of fluorescent colors to match instrument lasers and filters, and to accommodate co-staining with antibodies or other cell analysis reagents.
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Properties of cell trackers
| Short-term cell tracking | Long-term cell tracking |
|---|---|
| Rapid loading (typically, 15 min to 1 hr) | Typically, 1-hour cell loading |
| Short-term tracking (typically 3–6 generations or 72 hours) | Long-term tracking (typically 6–10 generations or 5–14 days) |
| Moderate resistance to photobleaching | High resistance to photobleaching |
| Moderate brightness | Intense brightness |
| Signal distribution throughout the cell | Punctate fluorescence within the cell |
| Multiplex easily with antibody labeling after fixing | Multiplex easily with antibody labeling |
Selection guide for cell trackers
| CellTracker dye | Common filter set | Ex/Em (nm) | Quantity | Cat. No. | |
|---|---|---|---|---|---|
| CellTracker Blue CMAC (7-amino-4-Chloromethylcoumarin) | CMAC | DAPI | 353/466 | 5 mg | C2110 |
| CellTracker Blue CMF2HC (4-chloromethyl-6,8-difluoro-7-hydroxycoumarin) | Blue CMF2HC | DAPI | 371/464 | 5 mg | C12881 |
| CellTracker Violet BMQC (2,3,6,7-tetrahydro-9- bromomethyl-1H,5H-quinolizino( 9,1-gh)coumarin) | Violet BMQC | DAPI | 415/516 | 5 x 0.1 mg | C10094 |
| CellTracker Green CMFDA (5-chloromethylfluorescein diacetate) | Green CMFDA | FITC | 492/517 | 20 x 50 µg | C7025 |
| CellTracker Orange CMRA | Orange CMRA | TRITC | 548/576 | 20 x 50 µg | C34551 |
| CellTracker CM-DiI | CM-DiI | TRITC | 553/570 | 20 x 50 µg | C7000 |
| CellTracker Red CMTPX | CMTPX | Rhodamine | 577/602 | 20 x 50 µg | C34552 |
| CellTracker Deep Red | Deep Red | Cy5 | 630/650 | 20 x 15 µg | C34565 |
Common filter set
|
Emission max (nm)
|
Quantity
| Cat. No. | |
|---|---|---|---|---|
| Qtracker 525 Cell Labeling Kit | GFP or FITC | 525 | 1 kit | Q25041MP |
| Qtracker 565 Cell Labeling Kit | 565 | 1 kit | Q25031MP | |
| Qtracker 585 Cell Labeling Kit | TRITC-A | 585 | 1 kit | Q25011MP |
| Qtracker 605 Cell Labeling Kit | 605 | 1 kit | Q25001MP | |
| Qtracker 625 Cell Labeling Kit | Texas Red | 625 | 1 kit | A10198 |
| Qtracker 655 Cell Labeling Kit | 655 | 1 kit | Q25021MP | |
| Qtracker 705 Cell Labeling Kit | Cy5 | 705 | 1 kit | Q25061MP |
| Qtracker 800 Cell Labeling Kit | Cy7 | 800 | 1 kit | Q25071MP |
| CellTracker dye | Common filter set | Ex/Em (nm) | Quantity | Cat. No. | |
|---|---|---|---|---|---|
| CellTracker Blue CMAC (7-amino-4-Chloromethylcoumarin) | CMAC | DAPI | 353/466 | 5 mg | C2110 |
| CellTracker Blue CMF2HC (4-chloromethyl-6,8-difluoro-7-hydroxycoumarin) | Blue CMF2HC | DAPI | 371/464 | 5 mg | C12881 |
| CellTracker Violet BMQC (2,3,6,7-tetrahydro-9- bromomethyl-1H,5H-quinolizino( 9,1-gh)coumarin) | Violet BMQC | DAPI | 415/516 | 5 x 0.1 mg | C10094 |
| CellTracker Green CMFDA (5-chloromethylfluorescein diacetate) | Green CMFDA | FITC | 492/517 | 20 x 50 µg | C7025 |
| CellTracker Orange CMRA | Orange CMRA | TRITC | 548/576 | 20 x 50 µg | C34551 |
| CellTracker CM-DiI | CM-DiI | TRITC | 553/570 | 20 x 50 µg | C7000 |
| CellTracker Red CMTPX | CMTPX | Rhodamine | 577/602 | 20 x 50 µg | C34552 |
| CellTracker Deep Red | Deep Red | Cy5 | 630/650 | 20 x 15 µg | C34565 |
Common filter set
|
Emission max (nm)
|
Quantity
| Cat. No. | |
|---|---|---|---|---|
| Qtracker 525 Cell Labeling Kit | GFP or FITC | 525 | 1 kit | Q25041MP |
| Qtracker 565 Cell Labeling Kit | 565 | 1 kit | Q25031MP | |
| Qtracker 585 Cell Labeling Kit | TRITC-A | 585 | 1 kit | Q25011MP |
| Qtracker 605 Cell Labeling Kit | 605 | 1 kit | Q25001MP | |
| Qtracker 625 Cell Labeling Kit | Texas Red | 625 | 1 kit | A10198 |
| Qtracker 655 Cell Labeling Kit | 655 | 1 kit | Q25021MP | |
| Qtracker 705 Cell Labeling Kit | Cy5 | 705 | 1 kit | Q25061MP |
| Qtracker 800 Cell Labeling Kit | Cy7 | 800 | 1 kit | Q25071MP |
CellTracker dyes
Cell tracker dyes provide the simplest processes for monitoring cell movement and localization. These dyes freely pass through the membrane of cells where they become cell-impermeant by reacting with thiol groups, except for CellTracker Deep Red which reacts with amine groups on proteins. Cell tracker dyes are passed to daughter cells for multiple generations but do not transfer to neighboring cells. They are stable, non-toxic at working concentrations (Figure 1), and brightly fluorescent at physiological pH (Figures 2, 3). CellTracker dyes have a wide range of emission spectra for multiplexing with antibody stains.
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Figure 1. CellTracker dyes do not contribute to cytotoxicity. HeLa cells were plated in 96-well plates at a density of 2,500 cells per well. The next day the cells (across three separate 96 well plates) were loaded with the indicated CellTracker at 1 µM concentration in Live Cell Imaging Solution (LCIS). DMSO in LCIS was applied to a column of cells for an internal control. Cells were incubated for 60 mins at 37°C and then loading media was aspirated, cells washed 1X in LCIS, and LCIS added back to wells for imaging. A 10% area of the well was imaged using the scan function of the EVOS FL Auto Imaging System, and then CyQUANT Direct Cell Proliferation Assay reagent was used to measure cell number. This experimental protocol was repeated for days 2 and 3 and represented as scanned and stitched area images and graphed to show cell proliferation assay results.
Figure 2. Live cell multiplexing using CellTracker Deep Red. HeLa cells were treated with CellTracker Deep Red (white), CellLight Mitochondrial-GFP, BacMam 2.0 (green), CellLight Golgi-RFP, BacMam 2.0 (red), and NucBlue Live ReadyProbes Reagent (blue).
Figure 3. CellTracker Deep Red can be used for fixed cell multiplexing. HeLa cells treated with CellTracker Deep Red (pink), CellLight Mitochondrial-GFP, BacMam 2.0 (green), anti-vimentin and goat-anti-rabbit Alexa Fluor 750 (white), and NucBlue Fixed Cell ReadyProbes Reagent (blue).
Qtracker labels
Qtracker labels provide longer-term tracking of cell movement and localization. These probes are more resistant to photobleaching and are retained through more cycles of cell division. Qtracker labels use a custom targeting peptide to be delivered to the cytoplasm of live cells, providing intense, stable signals for long-term (6–10 generations) tracking which does not transfer to adjacent cells. Qtracker labels offer eight distinct probe with different emissions from a single excitation for easy combination with antibody staining.
Figure 4. Multiplexing Qdot probes, dyes, and fluorescent proteins. U2-OS cells were transduced with CellLight Plasma Membrane-GFP (green), followed by labeling with Qtracker 655 Cell Labeling Kit (magenta), and HCS NuclearMask Blue (blue).
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