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CellBrite™ Fix Membrane Stains

CellBrite™ Fix Membrane Stains are a new class of fluorogenic membrane dyes that covalently stain the plasma membrane in live cells. They are unique among membrane stains in that they can withstand both fixation and detergent permeabilization. Available with green, red, and far-red fluorescence.

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Cellular localization

Membrane/cell surface, Membrane/vesicular

For live or fixed cells

For live/intact cells, Covalent & fixable stains

Assay type/options

Co-cultures, Long term staining (24-72h)

Fixation options

Fix after staining (methanol), Permeabilize after staining, Fix after staining (formaldehyde)

Colors

Green, Red, Far-red

Dye
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Set of 5 vials
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Product Description

CellBrite™ Fix Membrane Stains Features

  • The only membrane dyes that withstand fixation and permeabilization
  • Bright, uniform cell surface staining in 15 minutes
  • Available with green, red and far-red fluorescence
  • Each vial makes 20 uL of 1000X dye

CellBrite™ Fix dyes are a series of proprietary fluorophores developed by Biotium to rapidly stain the outer plasma membranes of live cells. Other membrane dyes like DiO, DiI, Vybrant® membrane dyes, CellMask™, CellVue®, or PKH dyes can be fixed with formaldehyde. But they are not compatible with detergent permeabilization or methanol fixation, because these treatments extract lipophilic dyes from membranes. In contrast, CellBrite™ Fix Membrane Stains are unique in that their surface staining can withstand permeabilization and methanol fixation, allowing plasma membrane staining to be combined with intracellular staining with antibodies. Unlike lectins such as WGA, which bind specific targets that may vary between cell types, CellBrite™ Fix dyes are general membrane stains.

CellBrite™ Fix dyes are amine-reactive and are designed to accumulate at the cell membrane, where they become covalently attached to membrane proteins. As a result, surface staining is well-retained after permeabilization or methanol fixation, with only a slight increase in intracellular fluorescence compared to formaldehyde fixation alone. CellBrite™ Fix dyes are only weakly fluorescent in aqueous media but become intensely fluorescent upon membrane staining. This fluorogenic property of the dyes makes the staining very specific with low background. Due to their better water solubility, CellBrite™ Fix dyes yield much more uniform staining compared to lipophilic carbocyanine dyes like DiO and DiI. The dyes are non-cytotoxic and do not readily transfer between cells. They also can be used to stain yeast and bacteria (gram-positive or gram-negative).

Because CellBrite™ Fix staining is covalent, the fluorophores remain on the outer leaflet of the cell membrane, unlike the traditional membrane dyes, such as DiO and DiI, which are known to undergo flip-flop transmembrane movement during staining. Thus, CellBrite™ Fix dyes may be uniquely suited for membrane potential detection when coupled with a quencher dye whose partition into the membrane is membrane-potential dependent, similar to the DiO/DPA system used for this purpose (see J Neurosci 29(29), 9197(2009)). We welcome any collaborator who may be interested in working with Biotium to validate CellBrite™ Fix dyes for this application to contact us at techsupport@biotium.com.

CellBrite™ Fix dyes must be used to stain live cells before fixation. They cannot be used to stain cells that are already fixed (the dyes primarily label intracellular membranes in fixed cells). Our original CellBrite™ Cytoplasmic Membrane Dyes can be used to stain cells after fixation. To find the right stain for your application, see our Membrane & Cell Surface Stains Comparison, or download our Membrane & Surface Stains Brochure.

Note that CellBrite™ Fix dye stain dead cell more intensely than live cells. With prolonged dye incubation, or if cells are cultured after staining, the dye also will be internalized by endocytosis, resulting in labeling of intracellular vesicles. Please see our Tech Tip: Five Steps for Success with Membrane and Surface Stains for tips on staining and imaging (step 5) with CellBrite™ Fix.

CellBrite™ Fix Membrane Stains are provided in lyophilized format. A vial of anhydrous DMSO is included for dye reconstitution. After reconstitution, each vial yields 20 uL of 1000X dye (download the product protocol for more information). The dyes are available with green, red, and far-red fluorescence. We also offer MemBrite™ Fix Cell Surface Staining Kits. with a wide selection of color options. For more information, see Frequently Asked Questions.

DyeAbs/EmSizeCatalog number
CellBrite™ Fix 488480/513 nmTrial size (1 vial)30090-T
Set of 5 vials30090
CellBrite™ Fix 555542/571 nmTrial size (1 vial)30088-T
Set of 5 vials30088
CellBrite™ Fix 640638/667 nmTrial size (1 vial)30089-T
Set of 5 vials30089

Vybrant® and CellMask™ are trademarks of Thermo Fisher Scientific. CellVue® is a registered trademark of Millipore Sigma.

 

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    LysoView™ Dyes

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    NucSpot® Live Cell Nuclear Stains

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    RedDot™2 Far-Red Nuclear Stain, 200X in DMSO

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Support & Faq

CellBrite™ & MemBrite™ Membrane & Cell Surface Stains

CellBrite™ Cytoplasmic Membrane Stains are lipophilic dyes for simple, non-toxic, stable labeling of membranes in live or fixed cells. Cells can be fixed with formaldehyde before or after CellBrite™ staining. But the staining has poor tolerance for permeabilization or methanol fixation, so CellBrite™ staining is not easily combined with intracellular immunofluorescence (IF) staining. The dyes also do not stain bacteria or yeast. CellBrite™ NIR dyes are CellBrite™ dyes with near-infrared fluorescence compatible with small animal NIR imaging systems.

CellBrite™ Fix and MemBrite™ Fix stains were developed to overcome some of these shortcomings. They are novel covalent stains that can be fixed and permeabilized for IF staining. CellBrite™ Fix Membrane Stains are fluorogenic reactive membrane dyes that rapidly accumulate at the plasma membrane. When they incorporate into lipids, they become fluorescent, and at the same time react covalently with membrane proteins for stable labeling. Staining takes only 15 minutes in a single step with no wash. CellBrite™ Fix stains mammalian cells, yeast, and bacteria.

MemBrite™ Fix Cell Surface Stains do not bind lipids, but label cell surface proteins. MemBrite™ Fix requires a two-step staining protocol with washing, but offers a more extensive choice of dye colors than CellBrite™ Fix. MemBrite™ Fix also can be used to stain yeast. But unlike original CellBrite™ dyes and lectins, CellBrite™ Fix and MemBrite™ Fix cannot be used on cells that are already fixed.

To select a dye that’s right for your application, see our Membrane and Cell Surface Stains Comparison, or download our Membrane & Surface Stains Brochure.

Category: CellBrite™ & MemBrite™ Membrane & Cell Surface Stains

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Original CellBrite™ Cytoplasmic Membrane Stains are lipophilic carbocyanine dyes. These dyes undergo an increase in fluorescence when they insert into lipid bilayers. Lipophilic carbocyanine dyes stably label the plasma membrane and other intracellular membranes of cells. They also can be used to stain fixed cells or artificial lipid bilayers. CellBrite™ cytoplasmic membrane stains are lipophilic carbocyanine dyes. These dyes undergo an increase in fluorescence when they insert into lipid bilayers. Lipophilic carbocyanine dyes stably label the plasma membrane and other intracellular membranes of cells. They also can be used to stain artificial lipid bilayers. Immediately after staining cultured cells, the dyes primarily localize to the plasma membrane. If cells are cultured over time after staining, the labeled membranes are internalized and staining gradually becomes mostly intracellular.

Category: CellBrite™ & MemBrite™ Membrane & Cell Surface Stains

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Lipophilic carbocyanine dyes like our original CellBrite™ Cytoplasmic Membrane Stains have been used to stain neuronal cells in culture for several weeks, and in vivo for up to a year (see note below). The dyes do not appreciably affect cell viability, and do not readily transfer between cells with intact membranes, allowing cell migration and tracking studies in mixed populations. Staining with the covalent stains CellBrite™ Fix and MemBrite™ Fix lasts up to 48 hours in tissue culture cells (see note below).

Note: Over time, all cell surface stains will be internalized and become intracellular as membranes turn over by endocytosis. The rate of internalization may vary by cell type, rate of membrane turnover, and rate of cell division. In immortalized cells in culture, most of the surface staining becomes internalized over the course of about 24 hours for CellBrite™, CellBrite™ Fix, and MemBrite™ Fix stains.

For an alternative stable and fixable stain for long term cell tracking or tracking cells in mixed cultures, see our ViaFluor® SE Cell Proliferation Dyes. These dyes covalently label intracellular proteins throughout the cell and are non-toxic and fixable.

Category: CellBrite™ & MemBrite™ Membrane & Cell Surface Stains

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Product shelf life, solubility, shipping and storage, and stability

Bioscience kits
The guaranteed shelf life from date of receipt for bioscience kits is listed on the product information sheet. Some kits have an expiration date printed on the kit box label, this is the guaranteed shelf life date calculated from the day that the product shipped from our facility. Kits often are functional for significantly longer than the guaranteed shelf life. If you have an older kit in storage that you wish to use, we recommend performing a small scale positive control experiment to confirm that the kit still works for your application before processing a large number of samples or precious samples.

Antibodies and other conjugates
The guaranteed shelf life from date of receipt for antibodies and conjugates is listed on the product information sheet. Antibodies and other conjugates often are functional for significantly longer than the guaranteed shelf life. If you have an older conjugate in storage that you wish to use, we recommend performing a small scale positive control experiment to confirm that the product still works for your application before processing a large number of samples or precious samples.

For lyophilized antibodies, we recommend reconstituting the antibody with glycerol and antimicrobial preservative like sodium azide for the longest shelf life (note that sodium azide is not compatible with HRP-conjugates).

Chemicals, dyes, and gel stains
Biotium guarantees the stability of chemicals, dyes, and gel stains for at least a year from the date you receive the product. However, the majority of these products are highly stable for many years, as long as they are stored as recommended. Storage conditions can be found on the product information sheet or product safety and data sheet, material safety data sheet, and on the product label. Fluorescent compounds should be protected from light for long term storage.

If you have a Biotium compound that has been in storage for longer than one year that you wish to use, we recommend performing a small scale positive control experiment to confirm that the compound still works for your application before processing a large number of samples or precious samples.

Expiration date based on date of manufacture (DOM)
If your institution requires you to document expiration date based on date of manufacture for reagents, please contact techsupport@biotium.com for assistance.

Chemical products with special stability considerations:

Esters

Ester compounds include the following:
• Succinimidyl esters (SE, also known as NHS esters), such as our amine-reactive dyes
• Acetoxymethyl esters (AM esters) such as our membrane-permeable ion indicator dyes
• Diacetate-modified dyes, like ViaFluor™ 405, CFDA, and CFDA-SE cell viability/cell proliferation dyes

Ester dyes are stable in solid form as long as they are protected from light and moisture. Esters are not stable in aqueous solution. Concentrated stock solutions should be prepared in anhydrous DMSO (see Biotium catalog no. 90082). Stock solutions in anhydrous DMSO can be stored desiccated at -20°C for one month or longer. Esters should be diluted in aqueous solution immediately before use. Succinimidyl esters (SE) should be dissolved in a solution that is free of amine-containing compounds like Tris, glycine, or protein, which will react with the SE functional group. AM esters and diacetate compounds should be dissolved in a solution that is free of serum, because serum could contain esterases that would hydrolyze the compound.

A note on CF™ dye succinimidyl ester stability
Succinimidyl esters are generally susceptible to hydrolysis, which can result in lower labeling efficiency. Heavily sulfonated dyes, such as the Alexa Fluor® dyes, DyLight® dyes and IRDyes® are particularly hygroscopic, worsening the hydrolysis problem. For example, the percent of active Alexa Fluor® 488 succinimidyl ester (SE) could be well below 50% by the time of application (according to the manufacturer’s product datasheet). In a number of Alexa Fluor® SE reactive dyes, the SE group is derived from an aromatic carboxylic acid, while in all of Biotium’s CF™ dyes the SE group is prepared from an aliphatic carboxylic acid. This structural difference reduces the susceptibility of CF™ dye SE reactive groups to hydrolysis, resulting in relatively stable reactive dyes with consistently higher labeling efficiency compared to other SE derivatives of other fluorescent dyes.

Maleimides, MTS and thiosulfate dyes
Like the succinimidyl ester dyes, these dyes are also susceptible to hydrolysis, although generally to a much lower degree. Thus, for long term storage, anhydrous DMSO is recommended for making stock solutions.

Other reactive dyes
Amines, aminooxy (also known as oxylamine), hydrazide, azide, alkyne, BCN, and tyramide reactive dyes, as well as dye free acids, are generally stable in aqueous solution when stored at -20°C for 6-12 months or longer, as long as no compounds are present that may react with the dye’s functional group. See the product information sheets for specific reactive dyes more information.

Coelenterazines and D-luciferin

Coelenterazines are stable in solid form when stored as recommended; they are not stable in aqueous solution. Concentrated coelenterazine stock solutions (typically 1-100 mg/mL) should be prepared in ethanol or methanol; do not use DMSO or DMF to dissolve coelenterazines, because these solvents will oxidize the compounds. Ethanol or methanol stocks of coelenterazine can be stored at -20°C or below for six months or longer; alcohol stocks may evaporate during storage, so use tightly sealing screw cap vials and wrap the vials with Parafilm for long term storage. Propylene glycol also can be used as a solvent to minimize evaporation. If the solvent evaporates, the coelenterazine will still be present in the vial, so note the volume in the vial prior to storage so that you can adjust the solvent volume to correct for evaporation if needed. Prepare working solutions in aqueous buffers immediately before use. Coelenterazines are stable for up to five hours in aqueous solution.

Aquaphile™ coelenterazines are water soluble formulations of coelenterazines. They are stable in solid form when stored as recommended. Aquaphile™ coelenterazines should be dissolved in aqueous solution immediately before use. They are stable for up to five hours in aqueous solution.

Note that coelenterazines are predominantly yellow solids, but may contain dark red or brown flecks. This does not affect product stability or performance. If your coelenterazine is uniformly brown, then it is oxidized and needs to be replaced.

D-luciferin is stable in solid form and as a concentrated stock solution when stored as recommended; it is not stable at dilute working concentrations in aqueous solution. Prepare concentrated D-luciferin stock solutions (typically 1-100 mg/mL) in water, and store in aliquots at -20°C or below for six months or longer. Prepare working solutions immediately before use.

Category: Mix-n-Stain™ Antibody Labeling Kits, DNA Quantitation Kits, NucView® Caspase 3 Enzyme Substrates, Luciferase Assays, CF® Dyes, Product shelf life, solubility, shipping and storage, and stability

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Many of our solid compounds are packaged by lyophilization, in which case they usually do not appear as fluffy powders, but form a film or coating on the sides of the vial. Simply add the appropriate volume of the recommended solvent to the vial to make the desired concentration stock solution, and swirl or gently vortex to mix. Make sure the solvent comes in contact with the inside walls of the vial to fully recover the product.

Category: Product shelf life, solubility, shipping and storage, and stability

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Most of our products are stable at room temperature for many days, but we recommend storage at 4°C or -20°C to prolong shelf life. In the case of many of our aqueous dye solutions, the compounds are very stable at room temperature, but we recommend cold storage to prevent the growth of mold or other microbes over time. Therefore, to save on shipping costs, products with recommended storage at 4°C or -20°C may ship at ambient temperature or with an ice pack. These products may thaw without affecting product performance. When you receive the product, place it under the recommended storage conditions.

Category: Mix-n-Stain™ Antibody Labeling Kits, DNA Quantitation Kits, NucView® Caspase 3 Enzyme Substrates, Luciferase Assays, CF® Dyes, Product shelf life, solubility, shipping and storage, and stability

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Shipping/Shelf-life

Shipping & Handling Disclaimer: Shipping and handling methods are calculated based on item temperature storage recommendation. Large, multi-item orders may be assessed additional fees. Shipping to Puerto Rico may be assessed additional fees. A Biotium customer service representative will contact you if additional fees are required. For expedited shipping, please request in the Order Notes section of the checkout page. Please note that products with recommended storage at 4°C or -20°C may ship at ambient temperature. This will not affect product performance. When you receive the product, place it under the recommended storage conditions.