Download BioLegend Chemical Probes

BioLegend Chemical Probes
Reagents to Boost Your Research
Chemical non-antibody probes can complement the use of antibody-based detection in
applications like cell health, proliferation, tracking and localization. This table can help
indicate which reagent is best suited for which application. Specifically, does the reagent
label live cells or only dead/ fixed cells? Can the reagent be used on tissue or only cells
in culture? Is the reagent suitable for microscopy and flow cytometry applications? Is
this probe able to be fixed with a paraformaldehyde-based fixative? Choose the reagent
that best suits the cell type and conditions of the assay.
BioLegend’s chemical probes provide the best prices, flexible sizes, and have been
validated for equivalent performance compared to other commercially available
products.
Cell Type
Suitability*
Non-Antibody Chemical Probes for Cellular Labeling
Chemical Probe
Alternative to:
MitoSpy™ Green
FM
MitoTracker®
MitoSpy™ Orange
CMTMRos
MitoSpy™ Red
CMXRos
Green FM
MitoTracker®
Orange
CMTMRos
MitoTracker®
Red CMXRos
Calcein-AM
Calcein Violet-AM
CellTrace™
Calcein
Violet-AM
CFDA-SE
Imaging Equivalent
Channel
Subcellular
Localization
Live
Alexa Fluor® 488,
FITC
Mitochondria
Alexa Fluor® 555, PE
Dead/
fixed
Sample
Suitability
Application
Fixation
Cells/
Culture
Flow
Microscopy
•
•
•
•
Mitochondria
•
•
•
•
•
Alexa Fluor® 594
Mitochondria
•
•
•
•
•
Alexa Fluor® 488,
FITC
Cytoplasm
•
•
•
•
BV421™
Cytoplasm
•
•
•
•
Alexa Fluor® 488,
FITC
Cytoplasm
•
•
•
•
•
•
•
•
•
•
•
•
Tissue
Tag-it Violet™
Proliferation
CellTrace™
Violet Cell
Proliferation
BV421™
Cytoplasm
Helix NP™ Blue**
SYTOX® Blue
Coumarin, BV421™
Nucleus
•
•
•
•
•
Helix NP™ Green**
SYTOX® Green
Alexa Fluor® 488,
FITC
Nucleus
•
•
•
•
•
Helix NP™ NIR**
To-Pro®-3
Alexa Fluor ® 647,
APC
Nucleus
•
•
•
•
•
CytoPhase™ Violet
DyeCycle®
Violet
Coumarin, BV421™
Nucleus
•
•
•
•
•
•
DRAQ5™
Alexa Fluor® 647, APC
Nucleus
•
•
•
•
•
•
DRAQ7™
Alexa Fluor® 700,
APC/Cy7
Nucleus
•
•
•
•
•
Retention with
PFA Treatment
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Toll-Free Tel: (US & Canada): 1.877.BIOLEGEND (246.5343)
Tel: 858.768.5800
biolegend.com
07-0123-00
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Cell Type
Suitability*
Non-Antibody Chemical Probes for Cellular Labeling
Chemical
Probe
Imaging Equivalent
Channel
Subcellular
Localization
7-AAD
PE/Cy5, PE/Dazzle™
594
Propidium
Iodide
DAPI
Alternative to:
Sample
Suitability
Application
Fixation
Dead/
fixed
Tissue
Cells/
Culture
Flow
Microscopy
Nucleus
•
•
•
•
•
PE/Cy5, PE/Dazzle™
594
Nucleus
•
•
•
•
•
Coumarin, BV421™
Nucleus
•
•
•
•
•
•
•
•
•
•
Live
Retention with
PFA Treatment
Zombie Dyes
Live/Dead
Live/Dead™
Fixable dyes
Several formats
available.
Cell Surface or
Cytoplasmic
Primary
Amines
Flash
Phalloidin™
Green 488**
Alexa Fluor®
488 Phalloidin
Alexa Fluor® 488
Actin
•
•
•
•
N/A
Flash
Phalloidin™
Red 594**
Alexa Fluor®
594 Phalloidin
Alexa Fluor® 594
Actin
•
•
•
•
N/A
Flash
Phalloidin™
NIR 647**
Alexa Fluor®
647 Phalloidin
Alexa Fluor® 647, APC
Actin
•
•
•
•
N/A
For more detailed information on all of these probes, please visit: biolegend.com/cell_health_proliferation
*If the probe can stain live cells, then the probe is cell membrane permeant, otherwise, it requires fixation for the cells to be dead, as the probe is cell membrane
impermeant.
**For Helix NP™ and Flash Phalloidin™ products, fixation and permeabilization are required for staining cells and tissue sections.
FAQs
What is the mechanism of binding for the MitoSpy™ dyes?
MitoSpy™ Green FM has an unknown mechanism of binding. Unlike MitoSpy™ Orange and Red, it is mitochondrial membrane potential
independent and the brightness of its staining is not indicative of cell health for that reason. MitoSpy™ Green FM also is not efficiently
retained with fixation, so its primary application is mitochondrial localization or density in live cells. MitoSpy™ Orange and Red, on the other
hand, will label mitochondria that are actively respiring and carry a negative redox potential across the inner leaflet of the mitochondrial
membrane. If the cell starts to enter necrosis or apoptosis, the cell’s metabolism and respiration will abate, causing the net polarization to
neutralize making it no longer an attractive target for binding of the MitoSpy™ Orange and Red.
There are so many probes for assessing live/dead status. Which one is right for me?
The most practical live/dead probe is an impermeant nucleic acid stain, like the Helix NP™ dyes, that enters a cell with a compromised
membrane. However, nucleic acid stains are not fixable, especially if the fixative contains any organic solvent. For applications where cells
will be fixed and additional intracellular staining will be done, the Zombie Live/Dead Fixable dyes are preferred. They label cells with a
compromised plasma membrane and will also covalently conjugate primary amine-containing proteins. Because this conjugation mechanism
isn’t conserved to only intracellular targets, each cell type must be titrated until the negative/ live peak overlays the unstained cells as much
as possible. If it is helpful to know not just that a cell has died, but to have a more subtle indication of vitality, Calcein-AM, Calcein VioletAM, CFDA-SE and Tag-it Violet™ Proliferation and Cell Tracking Probe are all fluorogenic esterase substrates. This means that healthy cells
with active metabolism will stongly convert the substrate to a fluorescent form. The dimmer the signal, the less healthy the cell is. Esterase
substrates and MitoSpy™ Orange and Red can be used with other markers of apoptosis like Annexin-V and impermeant nucleic acid stains to
more sensitively track a cell’s progress into cell health and death.
What are my options for cell cycle analysis?
If assessing cell cycle in live cells that will not be fixed, the options are limited to CytoPhase™ Violet and DRAQ5. If cells will be fixed and
permeabilized it is much easier to label with enough reagent to obtain stoichiometric binding. CytoPhase™ Violet, DRAQ5™, Helix NP™ Blue,
Helix NP™ NIR, DRAQ7™, propidium iodide and DAPI can all be used in this application.
Alexa Fluor® is a trademark of Life Technologies Corporation. Brilliant Violet™ is a trademark Sirigen Group Ltd. MitoTracker®, CellTrace™ Violet, SYTOX™, TO-PRO®,
Live/Dead™, and DyeCycle™ are trademarks of Thermo Fisher Scientific Inc. and its subsidiaries.
Toll-Free Tel: (US & Canada): 1.877.BIOLEGEND (246.5343)
Tel: 858.768.5800
biolegend.com
07-0123-00
World-Class Quality | Superior Customer Support | Outstanding Value