FLOW  CYTOMETERY REAGENTS

Bright, fixable dead cell stains, available in 12 colors
Non-toxic, fixable proliferation dyes designed as improvements to CFSE
Monitor apoptosis with the worlds only live-cell, real-time caspase-3 substrates
Dyes for monitoring mitochondrial membrane potential and apoptosis/necrosis status
Over 1000 monoclonal antibodies, and high-quality secondary antibodies, available in a wide array of dye conjugates
Buffers, fixation and permeabilization solutions, and more

Live-or-Dye™ Fixable Viability Staining Kits

Live-or-Dye™ fixable dead cell stains are cell membrane impermeable amine-reactive dyes. The dyes enter dead cells that have compromised membrane integrity and covalently label free amines on intracellular proteins (Fig. 1). Live-or-Dye™ labeling is extremely stable, allowing the cells to be fixed and permeabilized without loss of fluorescence or dye transfer between cells. The staining protocol has been optimized to maximize live/dead discrimination with minimal live cell staining, in order to prevent interference with immunostaining. Biotium offers a selection of 12 bright and photostable Live-or-Dye™ stains for maximal flexibility in multi-color analysis (Fig. 2). Our Live-or-Dye NucFix™ Red is a unique fixable nuclear-specific dead cell stain.

Live-or-Dye fixable viability stains
Figure 1. Principle of Live-or-Dye discrimination of live and dead cells.
Figure 2. Live-or-Dye™ kits are available with 12 bright and stable dye colors for multichannel flow cytometry.

To learn more about Live-or-Dye™ Fixable Viability Stains, please visit the Live-or-Dye™ technology page.

Catalog No.

Viability Dye

Laser line

Emission filter

Abs/Em Max

Applications

32002, 32002-T Live-or-Dye™ 350/448 355 nm DAPI or Violet 347/448 nm Flow Cytometry
32003, 32003-T Live-or-Dye™ 405/452 405 nm Pacific Blue® 408/452 nm Flow Cytometry
32009, 32009-T Live-or-Dye™ 405/545 405 nm AmCyan 395/545 nm Flow Cytometry
32004, 32004-T Live-or-Dye™ 488/515 488 nm FITC 490/515 nm Flow Cytometry,
Microscopy
32012, 32012-T Live-or-Dye™ 510/550 488 nm Spectral scan 516/549 nm Spectral Cytometry
32005, 32005-T Live-or-Dye™ 568/583 488 or 561 nm PE 562/583 nm Flow Cytometry,
Microscopy
32006, 32006-T Live-or-Dye™ 594/614 488 or 561 nm PE-Texas Red® 593/614 nm Flow Cytometry,
Microscopy
32007, 32007-T Live-or-Dye™ 640/662 633 or 640 nm APC 642/662 nm Flow Cytometry, Microscopy
32013, 32013-T Live-or-Dye™ 665/685 633 or 640 nm Spectral scan 667/685 nm Spectral Cytometry
32008, 32008-T Live-or-Dye™ 750/777 633 or 640 nm APC-Cy®7 755/777 nm Flow Cytometry
32011, 32011-T Live-or-Dye™ 787/808 785 or 808 nm APC-Cy®7 or IR800 783/803 nm Flow Cytometry
32010, 32010-T Live-or-Dye NucFix™ Red 488 or 532 nm PE-Texas Red® 520/593 nm Flow Cytometry,
Microscopy

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VIAFLUOR® SE CELL PROLIFERATION KITS

Cell Proliferation Monitoring by Flow Cytometry

Cell proliferation dyes diffuse passively into live cells and are used for long-term cell labeling. They are initially non-fluorescent esters of amine-reactive dyes, but are converted to fluorescent dyes by intracellular esterases. The dyes then covalently react with amine groups on intracellular proteins, forming fluorescent conjugates that are retained in the cell. Immediately after staining a single, bright fluorescent population will be detected by flow cytometry. Each cell division that occurs after labeling is revealed by the appearance of a successively dimmer fluorescent peak on a flow cytometry histogram (Figs. 3&4). Cell proliferation dyes can be used to track cell divisions in vivo or in vitro. The staining can withstand fixation and permeabilization for subsequent immunostaining.

ViaFluor® SE Cell Proliferation Kits

CFSE (also known as CFDA-SE or carboxyfluorescein diacetate, succinimidyl ester) has been used for many years to monitor cell proliferation by flow cytometry. While several alternatives have been developed in more recent years, CFSE remains popular and widely used. Biotium continues to offer CFSE under the tradename ViaFluor® CFSE. However, CFSE has several drawbacks including leakage from the cell, cell toxicity, and bleed-through into the PE and PE-TexasRed® channels.

ViaFluor® 405 SE and ViaFluor® 488 SE cell proliferation dyes were developed at Biotium to provide superior cell staining, fixability, and low toxicity. ViaFluor® 405 is excited with the violet laser and detected in the Pacific Blue® channel, and gives great peaks with no toxicity (Fig. 3). It behaves identially to the popular proliferation dye CellTrace™ Violet (Fig. 3). ViaFluor® 488 and ViaFluor® CFSE are both detected in the FITC channel, but ViaFluor® 488 was developed as a less toxic, less leaky and more fixable alternative to the classic dye CFSE (Fig. 4). It also has less bleed-through into other channels such as PE and PE-Texas-Red®.

Figure 3. Jurkat cells were stained with 5 uM cell proliferation dye: ViaFluor® 405 (left panel) or CellTrace™ Violet (right panel). Cells were analyzed in the Pacific Blue channel immediately after staining, and then every day for four days. Both dyes give sharp peaks showing the progressive decrease in signal caused by cell division.
Figure 4. Jurkat cells were stained with 3 uM cell proliferation dye: ViaFluor® 488 (left panel) or ViaFluor® CFSE (right panel). Cells were analyzed in the FITC channel immediately after staining, and then every day for three days. CFSE loses a lot of signal between initial staining and Day 1, limiting the number of cell divisions that can be measured, while ViaFluor® 488 retains dye better and gives nice sharp peaks for each division cycle.

To learn more about ViaFluor® Cell Proliferation Kits, please visit the ViaFluor® technology page.

Cell Division
Tracking Dyes

Catalog No.

Ex/Em (nm)

Flow detection
channel

Features

ViaFluor® 405 Cell Proliferation Kit 30068 408/452 Pacific Blue® • Track cell division by dye dilution using flow cytometry

• ViaFluor® 488 is a unique, improved green dye to replace CFSE

• ViaFluor® 405 replaces CellTrace™ Violet

ViaFluor® 488 Cell Proliferation Kit 30086 493/532 FITC
ViaFluor® CFSE Cell Proliferation Kit 30050 495/519 FITC

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NUCVIEW® CASPASE-3 SUBSTRATES

NucView® Caspase-3 Substrates for live cell apoptosis monitoring

NucView® Caspase-3 Substrates are novel fluorogenic substrates developed by Biotium to detect apoptosis in intact cells in real-time. Biotium has invented a new concept in the design of fluorogenic enzyme substrates. The enzyme substrate is attached to a fluorogenic DNA dye. Before cleavage, the dye is non-fluorescent and unable to bind to DNA. Upon enzymatic cleavage of the substrate, the dye is released and becomes capable of binding to DNA to emit fluorescence. In the case of NucView® Caspase-3 Substrates, the caspase-3/7 substrate peptide DEVD is attached to a DNA-binding dye. The substrate enters the cell cytoplasm where it is cleaved by caspase-3 in apoptotic cells to release the fluorogenic DNA dye, which stains the nucleus. NucView® can be fixed with formaldehyde after staining, but it cannot be used to stain fixed cells or tissues.

Features

  • Simple, homogenous assay for endpoint assay or real-time detection in live cells
  • Non-toxic, allowing for multi-day experiments
  • Detect caspase-3 activity and visualize apopototic nuclear morphology
  • For flow cytometry, fluorescence microscopy, or live cell imaging systems
  • Available as stand-alone substrates or in kits with other apoptosis or necrosis probes
  • Available with blue, green, or red fluorescence
  • NucView® 488 validated in more than 100 cell types and 200 publications – See the list
Figure 5. Principle of NucView® substrate technology. The substrate is non-fluorescent and does not bind DNA. After enzyme cleavage, the high-affinity DNA dye is released to bind DNA and become fluorescent.
Figure 6. NucView® 405 Caspase-3 Substrate fluorescence over time in staurosporine-treated Jurkat cells, analyzed in the Pacific Blue® channel of a BD LSRII flow cytometer.

To learn more about NucView® Caspase-3 substrates, and kits please visit the NucView® technology page.

NucView® Caspase-3 Substrates and Kits

Catalog No.

Features

NucView® 405 Caspase-3 Substrate, 1 mM in DMSO 10405 Blue fluorescence for flow cytometry in the Pacific Blue® channel or microscopy with the 405 nm laser
NucView® 405 Caspase-3 Substrate, 1 mM in PBS 10407 NucView® 405 substrate in PBS, for DMSO-sensitive cell types
NucView® 488 Caspase-3 Substrate, 1 mM in DMSO 10402 Green fluorescent substrate validated in more than 100 cell types and 200 publications
NucView® 488 Caspase-3 Substrate, 1 mM in PBS 10403 NucView® 488 substrate in PBS, for DMSO-sensitive cell types
NucView® 530 Caspase-3 Substrate, 1 mM in DMSO 10406 Orange fluorescence for microscopy in the Cy®3 channel or flow cytometry in the R-PE channel
NucView® 530 Caspase-3 Substrate, 1 mM in PBS 10408 NucView® 530 substrate in PBS, for DMSO-sensitive cell types
NucView® 488 and MitoView™ 633 Apotosis Detection Kit 30062 NucView® 488 and far-red fluorescent MitoView™ 633 for the Cy®5 channel
NucView® 488 and RedDot™ 2 Apoptosis & Necrosis Kit 30072 NucView® 488 and far-red dead cell DNA dye RedDot™ 2 for the Cy®5 channel
Dual Apoptosis Assay with NucView® 488 and CF®594 Annexin V 30067 NucView® 488 and red fluorescent Annexin V apoptosis probe
Dual Apoptosis Assay with NucView® 488 and CF®640R Annexin V 30073 NucView® 488 and far-red fluorescent Annexin V apoptosis probe

Please click on the link (cf-order-sheet) below to order from a wide of range of NucView® Caspase-3 substrates. Please email the form to sales@atlantisbioscience.com

Cf-order-sheet

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CELLULAR STAINS FOR FLOW

Mitochondrial Membrane Potential

Loss of mitochondrial membrane potential is a hallmark for apoptosis. Biotium offers MitoView™ 633 dye for membrane potential-sensitive staining of mitochondria by microscopy or flow cytometry. For simultaneous detection of mitochondrial membrane potential and caspase-3 activity, see the NucView®488 and MitoView™633 Apoptosis Assay.

We also offer MitoView™ 405, which changes localization upon mitochondrial depolarization, and MitoView™ Green, a membrane-potential independent mitochondrial dye that can be used to image mitochondria following mitochondrial depolarization, or after fixation.

In healthy cells, JC-1 dye aggregates in mitochondria as a function of membrane potential, resulting in red fluorescence with brightness proportional to the membrane potential. Conversely, in apoptotic and necrotic cells with diminished mitochondrial membrane potential, JC-1 exists in a green fluorescent monomeric form in the cytosol, allowing of cell viability to be assessed by measuring the ratio of red to green fluorescence by flow cytometry or fluorescence microplate reader.Rhodamine 123 is a green fluorescent mitochondrial membrane potential dye. Red fluorescent TMRM and TMRE are the preferred dyes for quantitative membrane potential measurements.

Figure 7. Timecourse of NucView®488 and MitoView™633 in staurosporine-treated Jurkat cells. As apoptosis progresses, MitoView™633 staining is lost and cells become positive for NucView®488.

Mitochondrial Dyes

Abs/Em

Detection channel

Potential-dependent?

Catalog no.

Size

MitoView™ 633 622/648 nm* Cy®5, APC* Yes 70055-T 50 ug
70055 20×50 ug
NucView® 488 & MitoView™ 633 Apoptosis Kit 500/530 nm (NucView®488)
622/648 nm* (MitoView­™ 633)
FITC/Cy®5* Yes 30062 100 assays
MitoView™ 405 398/440 nm DAPI Partial† 70070-T 50 ug
70070 20×50 ug
MitoView™ Green 490/523 nm FITC, GFP No 70054-T 50 ug
70054 20×50 ug
JC-1 Mitochondrial Membrane Potential Detection Kit 510/527 nm (cytoplasm)
585/590 nm (mitochondria)
FITC/Cy®3 Yes 30001 100 assays
JC-1 Chloride Salt 70011 5 mg
JC-1 Iodide Salt 70014 5 mg
Rhodamine 123 505/534 nm FITC Yes 70010 50 mg
TMRE 548/573 nm Cy®3 Yes 70016 25 mg
TMRE, 2 mM in DMSO 70016 0.5 mL
TMRM 548/573 nm Cy®3 Yes 70017 25 mg

Apoptosis/Necrosis Kits & Annexin V

Annexin V is a 36 kDa protein that has high affinity for phosphatidylserine (PS). During apoptosis, PS is translocated from the inner to the outer leaflet of the plasma membrane, where it can be stained by fluorescent conjugates of Annexin V. Annexin V conjugates are available with a wide selection of CF® dyes and other labels, including Near-IR CF® Dye Annexin V conjugates.

We offer many different staining kits with various combinations of dyes, including: NucView® caspase-3 substrate, our bright and photostable CF®488A Annexin V, a variety of dead cell nucleic acid stains, including propidium iodide (PI), 7-AAD, Ethidium Homodimer III (EthDIII), and RedDot™2. See the table below for more details on the various combination kits.

We offer dead cell nucleic acid stains separately as well, including far-red fluorescent RedDot™2 far-red dye and green fluorescent NucSpot® 470 stain.

Figure 8. Untreated and staurosporine-treated Jurkat cells stained with the Apoptotic, Necrotic and Healthy Cells Plus Kit. Apoptotic cells are stained green, necrotic/late apoptotic cells are stained red, and all cell nuclei are stained blue.

Apoptosis and Necrosis Staining Kits

Catalog No.

Features

CF®488A Annexin V and PI Apoptosis Kit 30061 • CF®488A stains apoptotic cells green for the FITC channel
• PI stains necrotic cells red for the Cy®3 or PE channels
• For microscopy or flow cytometry
CF®488A Annexin V and 7-AAD Apoptosis Kit 30060 • CF®488A stains apoptotic cells green for the FITC channel
• 7-AAD stains necrotic cells red for the PE-Cy®5 or FL3 channels
• For flow cytometry
Apoptosis & Necrosis Quantitation Kit Plus 30065 • CF®488A stains apoptotic cells green for the FITC channel
• EthD-III stains dead cells red for the Cy®3 or PE channels
• EthD-III has higher affinity and quantum yield than PI or 7-AAD
• For microscopy or flow cytometry
Apoptotic, Necrotic & Healthy Cells Quantitation Kit Plus 30066 • CF®488A stains apoptotic cells green for the FITC channel
• EthD-III stains dead cells red for the Cy® channel
• Hoechst stains all cell nuclei blue for the DAPI channel
• For microscopy
NucView®488 and RedDot™2 Apoptosis and Necrosis Kit 30072 • NucView®488 stains apoptotic nuclei green for the FITC channel
• RedDot™2 stains necrotic cells far-red for the Cy®5 or PE-Cy®5, APC, or FL3 channels
• For microscopy or flow cytometry
Annexin V Conjugates Multiple • Annexin V with our bright and photostable CF® dyes and a selection of other labels
Annexin V Conjugates, Azide-Free Multiple • Annexin V with our bright and photostable CF® dyes and a selection of other labels
• Lyophilized and preservative-free for real-time cell imaging
Annexin V Near-IR CF® Dye Conjugates Multiple • Annexin V conjugated to our superior Near-IR CF® dyes
• Lyophilized and preservative-free for in vivo use
Annexin Binding Buffer 99902 • Concentrated Annexin Binding Buffer for cell staining
NucSpot® 470 Nuclear Stain 40083 • Green fluorescent dead cell stain
• Also useful as a nuclear-specific counterstain for fixed cells
RedDot™2 Far-Red Nuclear Stain 40061 • Far-red nuclear stain for dead cells or fixed cells
• Detect in the Cy®5 channel
Ethidium homodimer III 40051 • Red fluorescent dead cell stain
• Higher affinity and quantum yield than PI
Propidium Iodide 40017 • Widely used red fluorescent dead cell stain
7-AAD 40037 • Fluorescent dead cell stain for the PE-Cy®5/FL3 channel
NucSpot® Far-Red 40085 • Designed as an improved replacement for 7-AAD
• Less bleed into the PE-Texas Red® channel compared to 7-AAD

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PRIMARY & SECONDARY ANTIBODY CONJUGATES

Primary Antibodies

  • Growing collection of more than 1000 monoclonal antibodies: View Flyer
  • Validated in IHC and other applications
  • Choose from 13 bright and photostable CF® dyes, R-PE, APC, PerCP, HRP, AP, or biotin
  • Isotype control antibodies available conjugated to the same dyes and labels
  • Available BSA-free, ready to use for Mix-n-Stain™ labeling
  • Affordable 100 uL sizes available
Figure 9. Flow cytometry analysis of Jurkat cells stained with CF®647 isotype (orange) or CF®647 anti-CD31 clone C31.7.

Secondary Antibodies

  • Wide selection of host and target species and anti-tag antibodies
  • Your choice of more than 20 CF® dye colors, R-PE, APC, HRP, AP or biotin
  • Highly cross-adsorbed, F(ab’)2 fragments, and isotype-specific options
  • Affordable 50 uL sizes available
Figure 10. Secondary antibody conjugates using CF® dyes give bright signal and low background, due to their improved water solubility over other dyes such as Alexa Fluor®. Here, the same goat anti-mouse antibody was conjugated with CF®633, Alexa Fluor® 647, or Alexa Fluor® 633. Jurkat cells were labeled with a mouse anti-CD3 antibody and then one of the secondary conjugates. The CF®633 conjugate gave the highest signal-to-noise due to low background and high specific signal.

To learn more about our primary and secondary antibodies, and kits please visit the Antibodies technology page.

Primary Antibodies – your choice of sizes and formats:

Search for Primary Antibodies

Format Concentration Size
CF® dye conjugates (13 colors) 0.1 mg/mL 100 uL or 500 uL
Biotin, HRP, or AP conjugates 0.1 mg/mL 100 uL or 500 uL
R-PE, APC, or Per-CP conjugates 0.1 mg/mL 250 uL
Purified, with BSA 0.2 mg/mL 100 uL or 500 uL
Purified, BSA-free (Mix-n-Stain™ Ready) 1 mg/mL 50 uL

Secondary Antibodies – available with many conjugations

Search for Secondary Antibodies

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BUFFERS, FIXATIVES, OTHER ACCESSORIES

We offer the following accessory products for your flow cytometry experiments:

Click here to see all of our flow cytometry accessory products.

Flow Cytometry Fixation/Permeabilization Kit