Cell Meter™ BX650 fixable viability dye

Detection of Jurkat cell viability by Cell Meter™ fixable viability dye. Jurkat cells were treated and stained with Cell Meter™ BX650 (Cat#22520), and then fixed in 3.7% formaldehyde and analyzed by flow cytometry.  The dead cell population (Blue peak)  is easily distinguished from the live cell population (Red peak)  with PerCP channel, and nearly identical results were obtained before and after fixation.

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Additional ordering information

Telephone	1-800-990-8053
Fax	1-800-609-2943
Email	sales@aatbio.com
Quotation	Request
International	See distributors
Shipping	Standard overnight for United States, inquire for international

Physical properties

Molecular weight	844.83
Solvent	DMSO

Spectral properties

Absorbance (nm)	518
Excitation (nm)	518
Emission (nm)	654

Storage, safety and handling

H-phrase	H303, H313, H333
Hazard symbol	XN
Intended use	Research Use Only (RUO)
R-phrase	R20, R21, R22
Storage	Freeze (< -15 °C); Minimize light exposure
UNSPSC	12171501

Overview SDS Protocol

Molecular weight

844.83

Absorbance (nm)

518

Excitation (nm)

518

Emission (nm)

654

Discrimination and exclusion of dead cells from live cells allows cleaner separation and identification of cell populations. Cell Meter™ fixable viability dyes are a large family of cell-impermeable fluorescent viability dyes that are optimized to match the major excitation lasers of common flow cytometers, such as 350, 405, 488, 633 and 647 nm. These dyes are impermeant to live cells but permeant to cells with compromised membranes. They irreversibly react with amine- and thiol-containing proteins and other cellular components. Since dead or fixed cells with a compromised membrane more readily react with Cell Meter™ fixable cell stains, thus stain brighter than live cells with an intact membrane, these dyes can be used to assess live vs. dead status of mammalian cells. There are a few factors to be considered when using these dyes, e.g., the titration of each dye to ensure that live cells have minimal to no staining. Cell Meter™ BX650 fixable cell stain is optimized to be excited with the blue laser at 488 nm with emission at 650 nm. Compared to other commercially similar viability dyes, this fixable viability dye is much more robust and stable.

Platform

Flow cytometer

Excitation	488 nm laser
Emission	695/40 nm filter
Instrument specification(s)	PerCP channel

Fluorescence microscope

Excitation	TexasRed filter set
Emission	TexasRed filter set
Recommended plate	Black wall/clear bottom

Example protocol

AT A GLANCE

Protocol summary

Prepare samples in HHBS (0.5 mL/assay)
Add Cell Meter™ BX650 to the cell suspension
Stain the cells at room temperature for 20 - 60 minutes
Wash the cells
Fix the cells (optional)
Examine the sample with flow cytometer using 695/40 nm filter (PerCP channel) and/or fluorescence microscope using Texas Red filter set

Important

Thaw at room temperature before starting the experiment.

PREPARATION OF STOCK SOLUTIONS

Unless otherwise noted, all unused stock solutions should be divided into single-use aliquots and stored at -20 °C after preparation. Avoid repeated freeze-thaw cycles.

Cell Meter™ BX650 stock solution (500X)

Add 200 µL of DMSO into the Cell Meter™ BX650 vial to make 500X stock solution.
Note The unused Cell Meter™ BX650 stock solution should be divided into single use aliquots and stored at -20°C. Avoid repeated freeze/thaw cycles.

SAMPLE EXPERIMENTAL PROTOCOL

Treat cells as desired.
Wash cells once with HHBS or the azide- and serum/protein-free buffer of your choice.
Resuspend cells at 5 - 10 × 10⁶/mL in HHBS or in the azide- and serum/protein-free buffer of your choice.
Add 1 µL of 500X Cell Meter™ BX650 stock solution to 0.5 mL of cells/assay and mix it well.
Incubate at room temperature, 5 % CO₂ incubator for 20 - 60 minutes, protected from light.
Note The optimal stain concentrations and incubation time should be experimentally determined for different cell lines.
Wash cells twice and resuspend cells with HHBS or the buffer of your choice.
Fix cells as desired (optional).
Analyze cells with a flow cytometer using 695/40 nm filter (PerCP channel) and/or fluorescence microscope using Texas Red filter set.

Calculators

Common stock solution preparation

Table 1. Volume of DMSO needed to reconstitute specific mass of Cell Meter™ BX650 fixable viability dye to given concentration. Note that volume is only for preparing stock solution. Refer to sample experimental protocol for appropriate experimental/physiological buffers.

	0.1 mg	0.5 mg	1 mg	5 mg	10 mg
1 mM	118.367 µL	591.835 µL	1.184 mL	5.918 mL	11.837 mL
5 mM	23.673 µL	118.367 µL	236.734 µL	1.184 mL	2.367 mL
10 mM	11.837 µL	59.184 µL	118.367 µL	591.835 µL	1.184 mL

Molarity calculator

Enter any two values (mass, volume, concentration) to calculate the third.

Mass (Calculate)		Molecular weight		Volume (Calculate)		Concentration (Calculate)		Moles
	/		=		x		=

Spectrum

Open in Advanced Spectrum Viewer

Spectral properties

Absorbance (nm)	518
Excitation (nm)	518
Emission (nm)	654

Product Family

Name	Excitation (nm)	Emission (nm)
Cell Meter™ BX520 fixable viability dye	491	516
Cell Meter™ BX590 fixable viability dye	492	579
Cell Meter™ IX830 fixable viability dye	811	822
Cell Meter™ RX660 fixable viability dye	649	664
Cell Meter™ RX700 fixable viability dye	690	713
Cell Meter™ RX780 fixable viability dye	629	767
Cell Meter™ VX450 fixable viability dye	406	445
Cell Meter™ VX500 fixable viability dye	433	498

Images

Figure 1. Detection of Jurkat cell viability by Cell Meter™ fixable viability dye. Jurkat cells were treated and stained with Cell Meter™ BX650 (Cat#22520), and then fixed in 3.7% formaldehyde and analyzed by flow cytometry. The dead cell population (Blue peak) is easily distinguished from the live cell population (Red peak) with PerCP channel, and nearly identical results were obtained before and after fixation.

References

View all 50 references: Citation Explorer

Fluorescence-based method is more accurate than counting-based methods for plotting growth curves of adherent cells.
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CFDA-SE Combined with MACSiBeads™ Particles to Evaluate the Inhibitory Effect of Treg Cells in vitro.
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Journal: Annals of clinical and laboratory science (2019): 740-747

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Journal: PloS one (2019): e0221878

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Journal: Cytometry. Part B, Clinical cytometry (2018): 363-368

Delineating the distinct role of AKT in mediating cell survival and proliferation induced by CD154 and IL-4/IL-21 in chronic lymphocytic leukemia.
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BFD-22 a new potential inhibitor of BRAF inhibits the metastasis of B16F10 melanoma cells and simultaneously increased the tumor immunogenicity.
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Journal: Toxicology and applied pharmacology (2016): 56-67