LysoTracker Deep Red |
Catalog No.GC27019
One-Click Copy Product Info
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Lysotracker probes are acidotropic fluorescent probes used to label and track acidic organelles within living cells. These probes possess several key features, including highly selective targeting of acidic organelles and efficient labeling of live cells at nanomolar concentrations.
Products are for research use only. Not for human use. We do not sell to patients.
Sample solution is provided at 25 µL, 10mM.
Lysotracker probes are acidotropic fluorescent probes used to label and track acidic organelles within living cells. These probes possess several key features, including highly selective targeting of acidic organelles and efficient labeling of live cells at nanomolar concentrations. Furthermore, the LysoTracker series is available in several fluorescent colors, making it particularly suitable for multicolor labeling experiments.
Lysotracker probes, consisting of a fluorescein linked to a weak base, are partially protonated at neutral pH, freely permeate cell membranes, and typically accumulate on spherical organelles. Studies have shown that Lysotracker probes must be used at extremely low concentrations (typically around 50 nM) to achieve excellent selectivity. While the retention mechanism of these probes is not fully understood, it is likely related to protonation and retention in acidic organelles, even though subsequent treatment of cells with weakly alkaline cell-permeable compounds generally does not reverse the staining. Studies of Lysotracker probe internalization kinetics have shown that dye uptake into live cells occurs at rates of only seconds. However, these lysosomal probes can alkalinize lysosomes; for example, prolonged incubation can induce an increase in lysosomal pH. Therefore, it is recommended to incubate cells with the probe for only 1-5 minutes before imaging.
LysoTracker Deep Red is a deep red fluorescent dye used to label and trace acidic organelles in living cells. It has maximum excitation/emission wavelengths of 647/668 nm and can be detected using the same filters as Cy5 or MitoTracker Deep Red. This product is stored in DMSO at a concentration of 1 mM. For lysosomal staining in living cells, the recommended working concentration is 50-75 nM, which should be adjusted based on the actual situation.
Instructions
Before use, remove the product and return it to room temperature. Centrifuge briefly at low speed to allow the lysate to settle to the bottom of the tube. For first-time use, aliquot and store in single-use quantities at -20°C, away from light and in a dry place. Avoid repeated freeze-thaw cycles. The optimal working concentration should be optimized based on experimental requirements, cell type, and cell or tissue membrane permeability.
1. Preparation of Working Solution
Dilute the 1 mM stock solution to the working concentration using growth medium or an appropriate buffer. The working solution should be prepared immediately before use. For the Lysotracker series of lysosomal probes, the recommended working solution concentration is 50-75 nM.
[Note]:
1) To reduce false positives caused by probe overloading, it is recommended to use as low a concentration as possible without compromising staining quality.
2) If cells are incubated in dye-free medium after staining, a decrease in fluorescence signal and vacuolization will be observed.
2. Staining Procedure (Adherent Cells)
1) Place cells on a coverslip in a culture dish and add appropriate medium to allow them to grow.
2) Once the cells have reached an appropriate density, aspirate the medium and add an appropriate amount of 37°C pre-warmed medium containing the probe (ensure that the cells are completely covered). Incubate the cells for 30 minutes to 2 hours under growth conditions appropriate for the specific cell type (the specific incubation time depends on the cell type).
3) Replace the staining solution with fresh medium without the probe and observe using a fluorescence microscope equipped with appropriate filters. If staining is insufficient, it is recommended to increase the dye concentration or extend the staining time to allow the dye to accumulate on lysosomes.
Note:
Kinetic studies of Lysotracker Green DND-26 internalization revealed that uptake of this probe by living cells is complete within seconds. However, a significant drawback is that this probe exhibits a lysosomal "alkalinization effect," resulting in an increase in lysosomal pH with prolonged incubation. Therefore, this probe is only useful as a pH indicator when cells are incubated at 37°C for 1-5 minutes.
3. Staining Procedure (Suspension Cells)
1) Pellet cells by centrifugation and remove supernatant.
2) Resuspend cells in an appropriate amount of probe-containing culture medium pre-warmed at 37°C and incubate for 30 minutes to 2 hours under growth conditions appropriate for the cell type of interest (the specific incubation time depends on the cell type).
3) Centrifuge, remove staining solution, and resuspend cells in fresh culture medium.
4) Observe using a fluorescence microscope equipped with appropriate filters. If staining is insufficient, it is recommended to increase the dye concentration or extend the staining time to allow the dye to accumulate on the lysosomes.
[Note]:
Alternatively, suspension cells may adhere to BD Cell-Tak-treated coverslips. In this case, staining can be performed using a method similar to that used for adherent cells.
Precautions
1) Protect from light throughout the entire procedure.
2) For your safety and well-being, please wear a lab coat and disposable gloves.
| Applications |
1) Appearance: Blue solution
2) Concentration: 1 mM in DMSO 3) Subcellular localization: Lysosomes 4) Color: Far-infrared 5) Detection instruments: Fluorescence microscope, fluorescence imager, microscope 6) Ex/Em: 647/668 nm |
| Shipping | Transport with ice packs. |
| Storage Conditions | Store at -20°C, protect from light |
| Usage | For research use only! Not for use on humans. |
Average Rating: 5 (Based on Reviews and 30 reference(s) in Google Scholar.)