MitoSpy™ Green FM

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Other Names
Mitochondrial labeling
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MitoSpy_Green_FM_IF_101415
HeLa cells were stained with 250 nM of MitoSpy™ Green FM (green) for 20 minutes and fixed with 4% paraformaldehyde (PFA) for ten minutes. Then the cells were stained with Alexa Fluor® 594 phalloidin for 20 minutes (red) and counterstained with DAPI (blue). The image was captured with a 60x objective.
  • MitoSpy_Green_FM_IF_101415
    HeLa cells were stained with 250 nM of MitoSpy™ Green FM (green) for 20 minutes and fixed with 4% paraformaldehyde (PFA) for ten minutes. Then the cells were stained with Alexa Fluor® 594 phalloidin for 20 minutes (red) and counterstained with DAPI (blue). The image was captured with a 60x objective.
  • MitoSpy_Green_FM_IF_2_101415
    HeLa cells were treated with 400 nM MitoSpy™ Green FM (Green) for 30 minutes, fixed with 4% paraformaldehyde (PFA) for fifteen minutes, permeabilized with 0.5% Triton X-100 for three minutes, and blocked with 5% FBS for 60 minutes. Then the cells were intracellularly stained with purified anti-Cytochrome c antibody (clone 7H8.2C12) overnight at 4°C followed by Alexa Fluor® 594 (Red) goat anti-mouse IgG for one hour at room temperature (Cat. No. 405326, 1:250 dilution, 2 µg/ml). Nuclei were counterstained with DAPI (Blue, Cat. No. 422801). The image was captured with a 60X objective using KEYENCE BZ-X700 fluorescence microscope. Exposure time (in seconds) is 1/20.
See high resolution ICC data and chemical structure...
Cat # Size Price Save
424805 5 x 50 µg ¥13,000
424806 20 x 50 µg ¥42,000
Description

MitoSpy™ mitochondrial localization probes are cell-permeant, fluorogenic chemical reagents that are used for labeling mitochondria of living cells. MitoSpy™ Green FM's attraction to the mitochondria is not based on membrane potential and thus can be used to measure mitochondrial mass of individual cells in flow cytometry.

Product Details
Technical data sheet

Product Details

Molecular Mass
671.88 g/mol
Preparation
The stock solution for MitoSpy™ Green FM is prepared by dissolving the lyophilized probe in dimethyl sulfoxide (DMSO) to make a final concentration of 1 mM by adding 74 µl of DMSO to each vial.
Storage & Handling
Store MitoSpy™ Green FM at -20°C.
Application

ICC - Quality tested

Recommended Usage

Each lot of this reagent is quality control tested by immunocytochemistry staining. For immunocytochemistry microscopy, a concentration range of 50nM to 500nM is recommended. It is recommended that the reagent be titrated for optimal performance for each application.

Application Notes

MitoSpy™ Green FM is excited at 490nm and emits at 516 nm.

1. Prior to reconstitution, spin down the vial of lyophilized reagent in a microcentrofuge to ensure the reagent is at the bottom of the vial.

2. Reconstitute MitoSpy™ Green FM to a 1 mM concentration with DMSO by adding 74 µl DMSO to an individual vial of lyophilized material. Protect the stock solution from light and keep frozen for storage.

3. Prepare the working solution for MitoSpy™ Green FM in 37°C culture medium (incomplete), this will vary by cell line and type of imaging required. 

     • If labeling mitochondria for live cell imaging, a concentration between 50 - 250 nM is recommended.
     • If cells are labeled live and then subsequently fixed, a concentration between 250 - 500 nM is recommended.

4. Grow cells to a desired confluency and wash once with warm 1 x PBS.

5. Add the diluted MitoSpy™ Green FM solution to the live cells and place in the 37°C incubator for 20-30 minutes.

6. Wash the cells twice with warm 1 X PBS or culture media.

7. If the cells will be imaged live, they can now be imaged with a fluorescence microscope.

If the cells need to be fixed:

     A. Fix the cells with 2-4% paraformaldehyde (PFA) for ten minutes at room temperature.
     B. Wash the cells twice with 1 x PBS.
     C. Regular IF staining protocol can be used for antibodies or other probe co-stains.

Antigen Details

Distribution

Mitochondria.

Biology Area
Apoptosis/Tumor Suppressors/Cell Death, Cell Biology, Mitochondrial Function, Neuroscience
Molecular Family
Mitochondrial Markers
Gene ID
NA

Related FAQs

There are no FAQs for this product.
Go To Top Version: 1    Revision Date: 10/19/2015

For research use only. Not for diagnostic use. Not for resale. BioLegend will not be held responsible for patent infringement or other violations that may occur with the use of our products.

 

*These products may be covered by one or more Limited Use Label Licenses (see the BioLegend Catalog or our website, www.biolegend.com/ordering#license). BioLegend products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products, reverse engineer functionally similar materials, or to provide a service to third parties without written approval of BioLegend. By use of these products you accept the terms and conditions of all applicable Limited Use Label Licenses. Unless otherwise indicated, these products are for research use only and are not intended for human or animal diagnostic, therapeutic or commercial use.

 

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