Confocal immunofluorescent analysis of HeLa cells using MitoTracker® Deep Red FM (blue pseudocolor), Vimentin (D21H3) XP® Rabbit mAb (Alexa Fluor® 555 Conjugate) #9855 (red), and Lamin A/C (4C11) Mouse mAb (Alexa Fluor® 488 Conjugate) #8617 (green).
Each vial of MitoTracker® Deep Red FM contains 50 μg lyophilized solid. To make a 1 mM stock solution, reconstitute the solid in 91.98 μl high quality DMSO. While the optimal staining concentration and incubation times can vary by application, typical results can be obtained by diluting the stock solution directly in growth media to a final concentration of 500 nM and incubating for 30 min at 37ºC. After incubation, fix the cells in ice-cold, 100% methanol for 15 min at -20ºC and rinse 3 times with PBS for 5 min. Proceed with immunostaining.
Store lyophilized solid at -20°C dessicated and protected from light. In lyophilized form, this reagent is stable for 12 months. Once reconstituted in DMSO, the solution can be stored at -20°C, protected from light and must be used within 2 weeks. Avoid freeze-thaw cycles.
MitoTracker® Deep Red FM is well retained after fixation allowing for further sample processing and immunostaining. Excitation: 644 nm, Emission: 665 nm, Molecular Weight: 543.58 g/mol
MitoTracker® dyes are cell permeable probes that contain a mildly thiol-reactive chloromethyl moiety for mitochondrial labeling. To label mitochondria, cells are simply incubated with MitoTracker® probes, which passively diffuse across the plasma membrane and accumulate in active mitochondria.
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
Alexa Fluor is a registered trademark of Life Technologies Corporation.
MitoTracker is a registered trademark of Molecular Probes, Inc.