Actively helping customers, employees and the global community during the coronavirus SARS-CoV-2 outbreak.  Learn more >>

Cell Navigator® Mitochondrion Staining Kit *Orange Fluorescence with 405 nm Excitation*

Image of HeLa cells stained with Cell Navigator® Mitochondrion Staining Kit in a Costar black wall/clear bottom 96-well plate.
Image of HeLa cells stained with Cell Navigator® Mitochondrion Staining Kit in a Costar black wall/clear bottom 96-well plate.
Ordering information
Price ()
Catalog Number22673
Unit Size
Find Distributor
Additional ordering information
Telephone1-408-733-1055
Fax1-408-733-1304
Emailsales@aatbio.com
InternationalSee distributors
ShippingStandard overnight for United States, inquire for international
Spectral properties
Excitation (nm)425
Emission (nm)522
Storage, safety and handling
H-phraseH303, H313, H333
Hazard symbolXN
Intended useResearch Use Only (RUO)
R-phraseR20, R21, R22
UNSPSC12352200

OverviewpdfSDSpdfProtocol


Excitation (nm)
425
Emission (nm)
522
Our Cell Navigator® fluorescence imaging kits are a set of fluorescence imaging tools for labeling sub-cellular organelles such as membranes, lysosomes, mitochondria, nuclei, etc. The selective labeling of live cell compartments provides a powerful method for studying cellular events in a spatial and temporal context. This particular kit is designed to label mitochondria of live cells in orange fluorescence of large Stokes Shift. The kit uses a proprietary dye that selectively accumulates in mitochondria probably via the mitochondrial membrane potential gradient. The mitochondrial indicator, a hydrophobic compound, easily permeates intact live cells and trapped in mitochondria after it gets into cells. This fluorescent mitochondrial indicator is retained in mitochondria for a long time since it carries a cell-retaining group. This key feature significantly increases the staining efficiency. The mitochondrial stain used in the kit is quite photostable, making the fluorescence quite robust. The kit provides all the essential components with an optimized cell-labeling protocol. It is suitable for proliferating and non-proliferating cells, and can be used for both suspension and adherent cells.

Platform


Fluorescence microscope

ExcitationViolet filter
EmissionViolet filter
Recommended plateBlack wall/clear bottom

Components


Component A: MitoViolet™ 5501 vial (100 µL, 500X DMSO stock solution)
Component B: Live Cell Staining Buffer1 bottle (50 mL)

Example protocol


AT A GLANCE

Protocol summary

  1. Prepare cells
  2. Add MitoViolet™ 550 working solution
  3. Incubate at 37°C for 30 minutes to 2 hours
  4. Wash and Replace with growth medium
  5. Analyze the cells under fluorescence microscope at Ex/Em = 405/550 nm (Violet filter set)

Important notes
Thaw all the components at room temperature before starting the experiment.

PREPARATION OF WORKING SOLUTION

Add 20 µL of 500X MitoViolet™ 550 (Component A) into 10 mL of Live Cell Staining Buffer (Component B) to make MitoViolet™ 550 working solution. Protect from light. Note: 20 µL of 500X MitoViolet™ 550 (Component A) is enough for one 96-well plate. The optimal concentration of the fluorescent mitochondrial indicator varies depending on the specific application. The staining conditions may be modified according to the particular cell type and the permeability of the cells or tissues to the probe.

For guidelines on cell sample preparation, please visit
https://www.aatbio.com/resources/guides/cell-sample-preparation.html

SAMPLE EXPERIMENTAL PROTOCOL

For adherent cells:

  1. Grow cells either in a 96-well black wall/clear bottom plate (100 µL/well/96-well plate) or on cover-slips inside a petri dish filled with the appropriate culture medium.

  2. When cells reach the desired confluence, add equal volume of MitoViolet™ 550 working solution.

  3. Incubate the cells in a 37°C, 5% CO2 incubator for 30 minutes to 2 hours.

  4. Wash and replace MitoViolet™ 550 working solution with Hanks and 20 mM Hepes buffer (HH buffer) or buffer of your choice (e.g. the buffer with growth medium at 1:1 concentration).

  5. Observe the cells using a fluorescence microscope with Violet filter set (Ex/Em = 405/550 nm). Note: It is recommended to increase either the labeling concentration or the incubation time to allow the dye to accumulate if the cells do not appear to be sufficiently stained.

For suspension cells:

  1. Centrifuge the cells at 1000 rpm for 5 minutes to obtain a cell pellet and aspirate the supernatant.

  2. Resuspend the cell pellets gently in pre-warmed (37°C) growth medium, and add equal volume of MitoViolet™ 550 working solution.

  3. Incubate the cells in a 37°C, 5% CO2 incubator for 30 minutes to 2 hours.

  4. Wash and replace MitoViolet™ 550 working solution with Hanks and 20 mM Hepes buffer (HH buffer) or buffer of your choice (e.g. the buffer with growth medium at 1:1 concentration).

  5. Observe the cells using a fluorescence microscope with Violet filter set (Ex/Em = 405/550 nm). Note: It is recommended to increase either the labeling concentration or the incubation time to allow the dye to accumulate if the cells do not appear to be sufficiently stained. Suspension cells may be attached to cover-slips that have been treated with BD Cell-Tak® (BD Biosciences) and stained as adherent cells.

Spectrum


Open in Advanced Spectrum Viewer
spectrum

Spectral properties

Excitation (nm)425
Emission (nm)522

Citations


View all 5 citations: Citation Explorer
Co-delivery of VP-16 and Bcl-2-targeted antisense on PEG-grafted oMWCNTs for synergistic in vitro anti-cancer effects in non-small and small cell lung cancer
Authors: Heger, Zbynek and Polanska, Hana and Krizkova, Sona and Balvan, Jan and Raudenska, Martina and Dostalova, Simona and Moulick, Amitava and Masarik, Michal and Adam, Vojtech
Journal: Colloids and Surfaces B: Biointerfaces (2017): 131--140
Inhibition of heme oxygenase-1 enhances the chemosensitivity of laryngeal squamous cell cancer Hep-2 cells to cisplatin
Authors: Lv, Xin and Song, Dong-mei and Niu, Ying-hao and Wang, Bao-shan
Journal: Apoptosis (2016): 489--501
Effective two-photon excited photodynamic therapy of xenograft tumors sensitized by water-soluble bis (arylidene) cycloalkanone photosensitizers
Authors: Zou, Qianli and Zhao, Hongyou and Zhao, Yuxia and Fang, Yanyan and Chen, Defu and Ren, Jie and Wang, Xiaopu and Wang, Ying and Gu, Ying and Wu, Feipeng
Journal: Journal of medicinal chemistry (2015): 7949--7958
Melatonin promotes adipogenesis and mitochondrial biogenesis in 3T3-L1 preadipocytes
Authors: Kato, Hisashi and Tanaka, Goki and Masuda, Shinya and Ogasawara, Junetsu and Sakurai, Takuya and Kizaki, Takako and Ohno, Hideki and Izawa, Tetsuya
Journal: Journal of Pineal Research (2015): 267--275

References


View all 70 references: Citation Explorer
Quantification of carbonylated proteins in rat skeletal muscle mitochondria using capillary sieving electrophoresis with laser-induced fluorescence detection
Authors: Feng J, Arriaga EA.
Journal: Electrophoresis (2008): 475
Calcium, mitochondria and apoptosis studied by fluorescence measurements
Authors: Roy SS, Hajnoczky G.
Journal: Methods (2008): 213
Fluorescence imaging of mitochondria in yeast
Authors: Swayne TC, Gay AC, Pon LA.
Journal: Methods Mol Biol (2007): 433
A fluorescence assay for peptide translocation into mitochondria
Authors: Martinez-Caballero S, Peixoto PM, Kinnally KW, Campo ML.
Journal: Anal Biochem (2007): 76
Fast electrophoretic analysis of individual mitochondria using microchip capillary electrophoresis with laser induced fluorescence detection
Authors: Duffy CF, MacCraith B, Diamond D, O'Kennedy R, Arriaga EA.
Journal: Lab Chip (2006): 1007
Discrimination of depolarized from polarized mitochondria by confocal fluorescence resonance energy transfer
Authors: Elmore SP, Nishimura Y, Qian T, Herman B, Lemasters JJ.
Journal: Arch Biochem Biophys (2004): 145
A fluorescence-based technique for screening compounds that protect against damage to brain mitochondria
Authors: Kristian T, Fiskum G.
Journal: Brain Res Brain Res Protoc (2004): 176
Determination of the cardiolipin content of individual mitochondria by capillary electrophoresis with laser-induced fluorescence detection
Authors: Fuller KM, Duffy CF, Arriaga EA.
Journal: Electrophoresis (2002): 1571
Fluorescence imaging of metabolic responses in single mitochondria
Authors: Nakayama S, Sakuyama T, Mitaku S, Ohta Y.
Journal: Biochem Biophys Res Commun (2002): 23
Visualisation of mitochondria in living neurons with single- and two-photon fluorescence laser microscopy
Authors: Dedov VN, Cox GC, Roufogalis BD.
Journal: Micron (2001): 653