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Arcturus™ Paradise™ PLUS 2 Round Kit, amino allyl, no solvents (Applied Biosystems™)

The ARCTURUS® Paradise® PLUS Reagent System permits unprecedented gene expression analysis of millions of previously inaccessible samples. It is an integrated solution for gene expression studies from formalin-fixed, paraffin-embedded (FFPE) tissues, including reagents for tissue staining, RNA isolation and RNA amplification. Amplify RNA from as little as 5 ng of fixed RNA using methods that have been specially optimized to overcome the challenges of cross-linked templates.

This kit provides reagents for 2-round in-vitro transcription, and aminoallyl aRNA labeling.

Key product features:
• Accelerated discovery process – unlock the tissue archives to access previously inaccessible samples
• Simplified procedure – achieve unprecedented results in six steps
• Streamlined application – integrate seamlessly into current workflow
• Efficient workflow – secure downstream success with the Paradise® QC Kit
• Comprehensive system – benefit from the only complete system for formalin-fixed samples

Unlock the Tissue Archives
Currently, tens of millions of tissue samples with known outcomes are available in tissue archives. The Paradise® PLUS Reagent System enables microarray and qRT-PCR analysis of these previously inaccessible samples, allowing retrospective studies to accelerate discoveries.

Achieve Unprecedented Results in Six Steps
Only six steps are required to take you from sample to results (See Figure 1).

Integrate Seamlessly into Current Workflow
The Paradise® PLUS Reagent System is compatible with all major microarray platforms. Robust and reproducible results are achieved using tools and techniques commonly employed in today’s research laboratories (See Figures 2, 3, 4, and 5).

Secure Downstream Success with the Paradise® QC Kit
Not all FFPE samples contain high quality RNA. Identify the best samples for your studies using a simple, real-time PCR research technique, and then proceed with confidence using these pre-qualified samples (See Figure 6).

Benefit from the Only Complete System for Formalin-fixed Samples
The Paradise® PLUS Reagent System is the only available solution that provides all the components necessary to prepare RNA from formalin-fixed, paraffin-embedded tissue samples for gene expression analysis. To ensure successful microarray results, the system includes reagents optimized for exceptional recovery of RNA and superior RNA amplification.

Integrated Systems for Microgenomics
The Paradise® PLUS Reagent System is a validated as part of the ARCTURUS® Complete System for Microgenomics®, an integrated solution for utilizing small quantities of RNA for gene expression analysis. This system features the ARCTURUSXT™ Laser Capture Microdissection (LCM) Instrument to procure pure cell populations. Arcturus Systems for Microgenomics enable accurate and sensitive microarray assays that reveal molecular signatures otherwise obscured in whole tissue samples or cell mixtures.

For Research Use Only. Not for use in diagnostics procedures.

pHrodo™ Red Phagocytosis Particle Labeling Kit for Flow Cytometry (Invitrogen™)

• Specifically detect phagocytosis and endocytosis with pH-sensitive fluorogenic dye - discriminate endcytosed from adherent and extracellular particles
• Reduced signal variability and improved timing in sensitive experiments - no need for wash steps or quencher dye
• Bright red fluorescence - conveniently multiplex with green dyes such as GFP, Fluo-4, or calcein

View a selection guide for all pHrodo Indicators used in imaging, flow cytometry and microplate assays.

The pHrodo™ Red dye gives faster and more accurate results than any other phagocytosis assay

The new Molecular Probes™ proprietary pH-sensitive rhodamine-based pHrodo™ Red dye is non-fluorescent at neutral pH, but turns bright red upon acidification. Because it is both fluorogenic and pH-sensitive, the pHrodo™ Red dye can be used as a specific sensor of phagocytic events; acidification of the phagosome following phagocytosis is marked by red fluorescence. It is therefore an ideal tool with which to study phagocytosis and its regulation by drugs and/or environmental factors.

Rapid and reproducible

Wash steps and quencher dyes are not needed since the pHrodo™ Red dye is non-fluorescent outside the cell, making the staining protocol simple and fast. The elimination of wash steps and quencher dyes also improves assay reproducibility, particularly in plate reader-based assays.

Flexible applications

Use ready-made pHrodo™ Red BioParticles™ conjugates for convenient analysis of phagocytosis of Gram-positive or Gram-negative bacteria, or the amine-reactive pHrodo™ Red SE form for labeling microorganisms or proteins of your choice.

Compatible with multiple platforms
pHrodo™ Red dye conjugates can be used in plate readers, fluorescence microscopy imaging, and flow cytometry applications. The pHrodo™ Red Phagocytosis Particle Labeling Kit and the pHrodo™ Red E. coli BioParticles™ Phagocytosis Kit for flow cytometry were designed for rapid and convenient measurements of phagocytic activity in whole blood samples by flow cytometry. The kits include all the reagents required for assessing particle ingestion and red blood cell lysis. The Labeling Kit also contains the reagents required for labeling microorganisms with the pHrodo™ Red dye. Sufficient reagents are provided in both kits for approximately 100 assays.

The optimal absorption and fluorescence emission maxima of the pHrodo™ Red BioParticles™ conjugate is approximately 560 nm and 585 nm, respectively. However, the fluorophore is readily excited with the 488 nm argon-ion laser installed on most flow cytometers.

CellTrace™ CFSE Cell Proliferation Kit, for flow cytometry (Invitrogen™)

CellTrace™ CFSE Cell Proliferation Kit is used for in vitro and in vivo labeling of cells to trace multiple generations using dye dilution by flow cytometry.

• Superior performance—bright, single-peak staining enables visualization of multiple generations
• Long-term signal stability—well-retained in cells for several days post stain
• Non-toxic—staining does not adversely effect cell health
• Simple, robust staining protocol

View a selection guide for all CellTrace™ Cell Proliferation Kits for flow cytometry.

Superior fluorescent staining
Successful proliferation analysis by dye dilution (see figure below) requires an extremely bright dye to distinguish fluorescently labeled cells from auto-fluorescence after several cell divisions. The intense fluorescent staining provided by CellTrace™ CFSE dye enables the visualization of eight or more generations of proliferating cells before the signal is overwhelmed by intrinsic cellular auto-fluorescence. Consistent, homogeneous staining results in very little fluorescence variation between cells in a population, so distinct generations can be seen without any requirement for complex modeling software.

Long-term signal retention
Unlike stains that label the lipid membrane of cells, CellTrace™ CFSE dye easily crosses the plasma membrane and covalently binds inside cells where the stable, well-retained fluorescent dye is designed to provide a consistent signal, even after several days in a cell culture environment.

Non-toxic
CellTrace™ CFSE dye binds covalently to all free amines on the surface and inside of cells and shows little cytotoxicity, with minimal observed effect on the proliferative ability or biology of cells. Researchers have used CFSE SE labeling to show that transplantable hematopoietic cells proliferate in vitro in response to stimulation by a growth factor cocktail.

Simple, robust staining protocol
The CellTrace™ CFSE Cell Proliferation Kit contains convenient single-use vials of dry dye to permit small-scale experiments without preparing excess quantities of dye. A stock solution is prepared by dissolving the contents of a vial in anhydrous DMSO prior to use. To stain 1 mL of cells in protein-free medium, 1 µL of this stock solution is typically used. Cells should be stained for 20 minutes at room temperature with gentle agitation. A brief wash with complete medium will then quench any dye remaining in solution.

Click-iT™ EdU Cell Proliferation Kit for Imaging, Alexa Fluor™ 555 dye (Invitrogen™)

The Click-iT EdU Cell Proliferation Kit for Imaging is a superior alternative to traditional proliferation assays that is optimized for fluorescence microscopy applications. In this assay the modified thymidine analogue EdU is efficiently incorporated into newly synthesized DNA and fluorescently labeled with a bright, photostable Alexa Fluor® dye in a fast, highly-specific click reaction. This fluorescent labeling of proliferating cells is accurate and compatible with antibody methods due to the mild click protocol.

• Simple—works the first time, every time, in less time
• Efficient—no denaturation steps or harsh treatment required
• Content-rich results—better preservation of cell morphology, antigen structure, and DNA integrity
• Consistent—not dependent on variable antibody lots for detection

The most accurate proliferation-detection methods are based on the incorporation and measurement of nucleoside analogues in newly synthesized DNA, with bromodeoxyuridine (BrdU) a commonly used analogue. BrdU-labeled DNA is quantitated using anti-BrdU antibodies following DNA denaturation by harsh methods (HCl, heat, or enzymes) to expose the BrdU molecules. This step is time consuming and difficult to perform consistently. The harsh treatment can also adversely effect sample integrity and quality, which makes co-staining with other antibodies challenging.

Superior Proliferation Methodology
The Click-iT EdU Cell Proliferation Kit for Imaging provides a superior alternative to BrdU assays for measuring cell proliferation. EdU (5-ethynyl-2'-deoxyuridine) is a nucleoside analog of thymidine and is incorporated into DNA during active DNA synthesis. With Click-iT® EdU, mild fixation and detergent permeabilization is sufficient for the small molecule-based Click-iT® EdU detection reagent to gain access to the DNA. As a consequence, the Click-iT® EdU imaging kit is not only easy to use, but more accurate and compatible with cell cycle analysis and other intracellular or extracellular targets for truly content-rich results.

This kit is optimized for fluorescence microscopy applications; visit the Click-iT® technology area of our website for kits designed for high-content imaging, fluorescence microplate, or flow cytometry platforms.

Learn more about Click-iT technology >
Find more tools for image-based detection of proliferating cells >

Notes:
The Click-iT® assay can be used on cells in culture or in vivo following EdU administration by feeding or injection methods.
The Click-iT® assay can be used with BrdU in dual pulse experiments by using the anti-BrdU (clone MoBu-1) antibody, which does not cross react with EdU.
The Click-iT® technology is compatible with immunohistochemical, immunocytochemical, and fluorescent dyes that are fixation tolerant or designed for fixed cell labeling.

pHrodo™ Green E. coli BioParticles™ Conjugate for Phagocytosis (Invitrogen™)

New pH-sensitive pHrodo® Green dye conjugates, like our pHrodo® Red dye conjugates, give faster and more accurate results than any other phagocytosis assay. pHrodo® Green conjugates are non-fluorescent outside the cell at neutral pH, but fluoresce brightly green at acidic pH such as in phagosomes. Get faster staining and more accurate results—without the need for wash steps or quencher dye.

• Specific detection of phagocytosis and endocytosis
• Reduced signal variability and improved timing in sensitive experiments
• Enough to do 100 tests when using 100 µL in each well of 96-well plate
• Multiplex with other compatible dyes, such as red florescent protein, TMRM, NucBlue™ Hoechst, and CellROX® Deep Red

The fluorescence of the novel pHrodo® Green dye dramatically increases as pH decreases from neutral to acidic, making it an ideal tool to study phagocytosis and its regulation by drugs and/or environmental factors. The lack of fluorescence outside the cell eliminates the need for wash steps and quencher dyes.

Use the ready-made pHrodo® Green E. coli BioParticles® Conjugate in imaging, high content screening, high throughput screening, and flow applications. pHrodo® Green dye is also available as a conjugate of zymosan BioParticles®, S. aureus BioParticles®, or dextran 10,000 MW. To create other conjugates, such as antibody conjugates, use pHrodo® Green STP ester or pHrodo® Green maleimide.

For Research Use Only. Not for human or animal therapeutic or diagnostic use.

eBioscience™ Annexin V Apoptosis Detection Set PE-Cyanine7 (Invitrogen™)

Annexins are a family of calcium-dependent phospholipid-binding proteins that preferentially bind phosphatidylserine (PS) in all mammalian species. Under normal physiologic conditions, PS is predominantly located in the inner leaflet of the plasma membrane. Upon initiation of apoptosis, PS loses its asymmetric distribution across the phospholipid bilayer and is translocated to the extracellular membrane leaflet marking cells as targets of phagocytosis. Once on the outer surface of the membrane, PS can be detected by fluorescently labeled Annexin V in a calcium-dependent manner.

In early-stage apoptosis, the plasma membrane excludes viability dyes such as propidium iodide (PI), 7-AAD, or Fixable Viability Dyes such as eFluor™ 660 or eFluor™ 780. These cells will stain with Annexin V but not a viability dye, thus distinguishing cells in early apoptosis. However, in late stage apoptosis, the cell membrane loses integrity thereby allowing Annexin V to also access PS in the interior of the cell. A viability dye can be used to resolve these late-stage apoptotic and necrotic cells (Annexin V, viability dye-positive) from the early-stage apoptotic cells (Annexin V positive, viability dye-negative).

Due to the emission spectrum of PE-Cyanine7, the Annexin V Apoptosis Detection Set PE-Cyanine7 is not compatible with propidium iodide and 7-AAD. It is recommended to substitute a Fixable Viability Dye such as eFluor™ 660 or eFluor™ 780 in their place.

Not included:
Fixable Viability Dye eFluor™ 660 (cat. 65-0864)
Fixable Viability Dye eFluor™ 780 (cat. 65-0865)

Note: Fixable Viability Dye eFluor™ 450 is not recommended for use with Annexin V Apoptosis Detection Kits.

Reactivity/Species
Human, Mouse, Rat

Reported Application
Flow Cytometric Analysis

CaspGLOW™ Fluorescein Active Caspase-3 Staining Kit (Invitrogen™)

The CaspGLOW™ Fluorescein Active Caspase-3 Staining Kit contains the reagents to sensitively detect active Caspase-3 in mammalian cells. This assay utilizes the inhibitor specific for Caspase-3, DEVD-FMK, that is directly conjugated to FITC (fluorescein isothiocyanate) for the detection system. This reagent is cell permeable, non-toxic and irreversibly binds to the active enzyme. Detection of the labeled cells can be determined by flow cytometry, fluorescent microscopy or a fluorescent plate reader.

The Caspases are a family of aspartate-specific cysteine proteases that act in a step-wise signaling manner like kinases. Recruitment of these proteases to oligomerized receptors leads to activation and autoproteolytic cleavage. Active caspases can proteolyze additional caspases generating a caspase cascade. The final outcome of this signaling pathway is a form of controlled cell death, termed apoptosis.

Caspase-3, also known as Yama, CPP32, and apopain, cleaves its substrates at the carboxyl terminus of aspartate residues. Active Caspase-3 consists of 2 sets of homodimers (~ 17 and 12 kDa) that are derived from two precursor Caspase-3 polypeptides and has two active sites. Caspase-3 is proteolytically activated by other caspases.

Both subunits contribute to substrate binding and catalysis. The active site cysteine that covalently binds the substrate is located near the C-terminus of the large subunit. Active Caspase-3 has two-fold symmetry, two active site pockets each residing on an opposite side. Caspase-3, together with Caspases 8 and 9, is situated at pivotal junctions in apoptotic pathways. Caspase-3 appears to amplify Caspase 8 and 9 initiation signals into complete commitment to apoptotic disassembly.

Reported Application
Flow Cytometric Analysis, Immunocytochemistry

APO BrdU (TUNEL) Kit (Invitrogen™)

The ApoTarget™ APO-BRDU Kit is a two-color staining method for labeling DNA breaks and total cellular DNA to detect apoptotic cells by flow cytometry.

View a selection guide for all apoptosis assays for flow cytometry.

LIVE/DEAD™ Cell Vitality Assay Kit, C12 Resazurin/SYTOX™ Green (Invitrogen™)

The LIVE/DEAD Cell Vitality Assay Kit provides a simple, two-color fluorescence assay that distinguishes metabolically active cells from injured cells and dead cells. The assay is based on the reduction of C12-resazurin to red-fluorescent C12-resorufin in metabolically active cells and on the uptake of the cell-impermeant, green-fluorescent nucleic acid stain, SYTOX Green dye, in cells with compromised plasma membranes (usually late apoptotic and necrotic cells). In this assay, dead cells emit mostly green fluorescence and healthy, metabolically active cells emit mostly red fluorescence; injured cells exhibit reduced red and green fluorescence.

View a selection guide for all Cell Viability Assays for Flow Cytometry.

eBioscience™ Mouse Regulatory T Cell Staining Kit #3 (Invitrogen™)

The FJK-16s antibody reacts with mouse/rat Foxp3 also known as FORKHEAD BOX P3, SCURFIN, and JM2; cross reactivity of this antibody to other proteins has not been determined. Foxp3 a 49-55 kDa protein, is a member of the forkhead/winged-helix family of transcriptional regulators, and was identified as the gene defective in ‘scurfy’ (sf) mice. Constitutive high expression of Foxp3 mRNA has been shown in CD4+CD25+ regulatory T cells (Treg cells), and ectopic expression of Foxp3 in CD4+CD25- cells imparts a Treg phenotype in these cells.

Immunoblotting with FJK-16s antibody has mapped the epitope to amino acids 75-125 of the mouse Foxp3 protein. In the human, this region has been shown to be alternatively spliced at the mRNA level. Both the alternatively-spliced and non-spliced isoforms are present in the CD4+CD25+ subset of lymphocytes. Preliminary RT-PCR experiments have not revealed this alternatively-spliced isoform in mouse splenocytes, suggesting different gene regulation in the mouse and human.

Intracellular staining of mouse splenocytes with FJK-16s using the mouse Foxp3 staining sets and protocol reveals approximately 2% of total splenocytes in the C57Bl/6 strain and approximately 3-5% in the BALB/c mouse strain. Multicolor flow cytometric analysis demonstrates approximately 90% of the CD4+CD25+ cells and 4% of the CD4+CD25- cells staining with FJK-16s. B220+, CD11b+, CD11c+, and Ly6G/Gr-1+ cells do not show significant co-staining with FJK-16s. These data are consistent with a recent report which follows expression of Foxp3 using a GFP knock-in (Fontenot et al, 2005).

FJK-16s cross-reacts with rat Foxp3 This has been demonstrated by intracellular staining of Foxp3 and flow cytometry of rat splenocytes using the same method and reagents as used for mouse tissue. Please note that the CD4 and CD25 antibodies included in this kit only recognize the mouse antigens. For staining rat tissue, please use (CD4 FITC cat. 11-0040 and CD25 PE cat. 12-0390)

The anti-mouse Foxp3 Staining Set has been formulated and optimized for the staining of mouse splenocytes with the FJK-16s monoclonal antibody.

Not included:
Isotype controls for anti-CD4 (rat IgG2a, cat. 11-4321) and anti-CD25 (rat IgG1, cat. 12-4301)

Host
Rat

Isotype
IgG2a

Reactivity/Species
Mouse

Reported Application
Intracellular Staining Followed by Flow Cytometric Analysis

CaspGLOW™ Fluorescein Active Caspase-9 Staining Kit (Invitrogen™)

The CaspGLOW™ Fluorescein Active Caspase-9 Staining Kit contains all the reagents necessary to detect active caspase-9 in mammalian cells with high sensitivity. Fluorescein (FITC)-conjugated LEHD-FMK, a specific inhibitor of caspase-9, is utilized in this assay for detection. This reagent is cell permeable, non-toxic and irreversibly binds to the active enzyme. Detection of the labeled cells can be determined by flow cytometry, fluorescent microscopy or a fluorescent plate reader.

The caspases constitute a family of aspartate-specific cysteine proteases that mediate a sequence of cleavage events. Recruitment of the inactive proenzyme to oligomerized receptors leads to caspase activation and autoproteolytic cleavage. These active enzymes can then cleave other caspases, thereby generating a caspase signaling cascade that leads to a form of programmed cell death termed apoptosis.

Caspase-9, which is also known as ICE-LAP6 and Mch6, cleaves its substrates at the C-terminal aspartic acid residue of the motif Asp-X-X-Asp. Activation of this signaling protease occurs upon stimulation of death receptors such as CD95/APO-1/Fas, TRAIL-R1, TRAIL-R2, TNFR1, and TRAMP. Caspase-9 is activated by recruitment and dimerization within the Apaf-1 apoptosome. Once recruited, caspase-9 undergoes proteolytic cleavage at Asp315 to yield 35-kDa and 10-kDa fragments. Unlike other caspases, this cleavage event is not required for the catalytic activity of caspase-9. As an initiator caspase, this protease initiates a caspase signaling cascade that results in apoptosis. Substrates of caspase-9 include caspases-3, -6, and -7. Finally, dimerization of cleaved caspase-9 is inhibited by X-linked inhibitor of apoptosis (XIAP).

Reported Application
Flow Cytometric Analysis, Immunocytochemistry

eBioscience™ BrdU Staining Kit for Flow Cytometry PE (Invitrogen™)

The eBioscience BrdU Staining Kit for Flow Cytometry contains the necessary reagents and buffers for identifying and examining proliferating cells of mammalian species by flow cytometric analysis. Cycling cells are incubated with 5-bromo-2’-deoxyuridine (BrdU), a synthetic analog of thymidine which incorporates into newly synthesized genomic DNA during the S-phase of mitosis. Following DNA denaturation, the cells are stained for BrdU incorporation along with any other cell surface and/or intracellular targets of interest.

This kit is optimized to achieve brighter staining compared to traditional protocols.

Reactivity/Species
Bovine, Canine, Cow, Human, Monkey, Mouse, Non-human primate, Rat, Rhesus

Conjugate
PE

Laser
Blue Laser

Emit
575 nm

Excite
488 nm

Reported Application
Intracellular Staining Followed by Flow Cytometric Analysis

EnzChek™ Caspase-3 Assay Kit #1, Z-DEVD-AMC substrate (Invitrogen™)

The EnzChek Caspase-3 Assay Kit #1 allows detection of apoptosis by providing a simple and reliable method for assaying caspase-3 activity. The basis for the assay is the aminomethylcoumarin (AMC)-derived substrate Z-DEVD-AMC. This substrate, which is weakly fluorescent in the UV range (excitation/emission maxima ~330/390 nm), yields a bright, blue-fluorescent product (excitation/emission maxima ~342/441 nm) upon proteolytic cleavage. The kit can be used to continuously measure the activity of caspase-3 and closely related proteases in cell extracts and purified enzyme preparations, using a fluorometer or fluorescence microplate reader.

LIVE/DEAD™ BacLight™ Bacterial Viability Kit, for microscopy & quantitative assays (Invitrogen™)

The LIVE/DEAD BacLight Bacterial Viability Kit *for microscopy and quantitative assays* is a convenient and easy-to-use kit for monitoring the viability of bacterial populations as a function of the membrane integrity of the cell. Cells with a compromised membrane that are considered to be dead or dying will stain red, whereas cells with an intact membrane will stain green.

View additional information about all microbiological analysis products.

Premo™ Autophagy Sensor GFP-p62 Kit (Invitrogen™)

The Premo™ Autophagy Sensor GFP-p62 Kit combines the ability to monitor the induction or inhibition of autophagy, through the localization of the autophagy receptor p62 (also known as SQSTM1), with the high transduction efficiency and minimal toxicity of BacMam 2.0 technology. To perform an image-based analysis for autophagy, simply add the BacMam 2.0 GFP-p62 reagent to your mammalian cells, incubate overnight to ensure adequate protein expression, and then visualize using standard GFP (green fluorescent protein) settings. The GFP gene included in this chimera is TagGFP2, which has been demonstrated to mature 1.6-times faster than TagGFP and has increased pH stability when compared to Emerald GFP.

Each Premo™ Autophagy Sensor GFP-p62 Kit includes BacMam 2.0 GFP-p62 reagent and chloroquine diphosphate, which is used to inhibit autophagy.

This GFP-p62 chimera is recommended for use with red-emitting fluorescent proteins or dyes.

Highly efficient delivery system: >90% transduction of a wide range of mammalian cell lines, including primary cells, stem cells, and neurons
Fast and convenient: simply add the Premo™ Autophagy reagent to your cells, incubate overnight, and image—or store frozen, assay-ready cells for later use
Robust: the Premo™ Autophagy reagents are non-replicating in mammalian cells, lack observable cytopathic effect, and are suitable for biosafety level (BSL) 1
Multiplex-enabled: additional BacMam or fluorescent reagents can be used in conjugation with the Premo™ Autophagy products to detect and monitor additional targets of interest

The p62 protein, also known as sequestosome (SQSTM1), is an ubitiquitin-binding protein that functions as a receptor for cargos destined to be degraded by the cellular autophagic machinery. When autophagy is induced the p62 protein localizes to the autophagosomes and is subsequently degraded. Conversely, with the inhibition of autophagy, the p62 protein accumulates in the autophagosome. Thus, the subcellular localization a p62-fluorescent protein chimera serves as a useful marker for the induction and inhibition of autophagy.

To ensure reliable, high levels of expression in a wide variety of mammalian cells, the Premo™ Autophagy Sensors utilize BacMam 2.0 technology. BacMam reagents (insect Baculovirus with a Mammalian promoter) do not replicate in mammalian cells, are non-cytoxic (biosafety level 1), and are ready to use. Unlike expression vectors, BacMam reagents enable titratable and reproducible expression and offer high co-transduction efficiency; therefore, multiple BacMam reagents can be used in the same cell.

Each Premo™ Autophagy Sensor GFP-p62 Kit includes a vial of chloroquine diphosphate. Chloroquine has been demonstrated to inhibit autophagy by elevating lysosomal pH and therefore inhibiting the fusion of autophagosomes with lysosomes and preventing the subsequent lysosomal protein degradation.

LIVE/DEAD™ Yeast Viability Kit (Invitrogen™)

The LIVE/DEAD® Yeast Viability Kit combines a novel two-color fluorescent probe for yeast viability, FUN® 1, with a fluorescent fungal surface labeling reagent Calcofluor® White M2R. Plasma membrane integrity and metabolic function of fungi are required to convert the yellow-green-fluorescent intracellular staining of FUN® 1 into red-orange intravacuolar structures; Calcofluor White M2R labels cell-wall chitin with blue-fluorescence regardless of metabolic state.

Click-iT™ EdU Colorimetric IHC Detection Kit (Invitrogen™)

The Click-iT™ EdU Colorimetric IHC (Immunohistochemistry) Detection Kit is a superior alternative to traditional methods for the colorimetric detection of proliferating cells within tissue sections. In this assay, 5-ethynyl-2'-deoxyuridine (EdU), a nucleoside analog of thymidine containing an alkyne moiety, is incorporated into newly synthesized DNA. When biotin containing an azide group is added, a brief "click reaction" attaches the azide to the alkyne moiety on the EdU molecule. Upon addition of streptavidin–peroxidase and a peroxidase substrate, an enzymatic reaction generates a signal that can be detected with light microscopy and stored for future analysis.

Find more tools for image-based detection of proliferating cells >

• Optimized—enables colorimetric detection of proliferating cells in either tissue or cell samples
• Efficient—no denaturation steps or harsh treatment required
• Robust—improved preservation of cell morphology and the cellular environment offers content-rich results
• Consistent—not dependent on variable antibody lots for detection
• Simple—designed to work consistently, in less time than traditional methods

Traditional methods yield inconsistent results
Measuring changes to cell proliferation is a fundamental method for assessing cell health, determining genotoxicity, and evaluating anti-cancer drug research. The most accurate method of doing this is by directly measuring DNA synthesis. The traditional method utilizes the nucleoside analog 5-bromo-2'-deoxyuridine (BrdU), a nucleoside analog of thymidine, which is incorporated into newly transcribed DNA. After incorporation, the samples are treated with harsh methods (HCl, heat, or enzymes) to denature the DNA and expose the BrdU molecules to detection by anti-BrdU antibodies. These harsh treatments can adversely affect sample integrity, cell morphology, and image quality. In addition, manufacturing variability, nonspecific binding, and cross-reactivity issues can result in inconsistent results.

Advantages over other assays
The Click-iT EdU Colorimetric IHC Detection Kit offers many advantages over traditional methods like the BrdU assay. The small size of the biotin-azide moiety allows efficient detection of incorporated EdU using mild conditions. In contrast, BrdU assays require harsh denaturation methods using HCl and/or heat-induced epitope retrieval (HIER). Additionally, unlike the BrdU assay, which relies on antibodies that can sometimes exhibit nonspecific binding, the Click-iT EdU Colorimetric IHC Detection Kit utilizes a bioorthogonal (biologically unique) moiety—resulting in low background signal, high detection sensitivities, and no cross-reactivity issues.

The kit contains all the components needed to detect incorporated EdU present in formalin-fixed, paraffin-embedded (FFPE) tissue samples. The kit includes sufficient reagents for labeling fifty (50) 18 x 18 mm coverslips using 100 μL reaction reagent per test.

Vybrant™ Phagocytosis Assay Kit (Invitrogen™)

The Vybrant® Phagocytosis Assay Kit provides a way for researchers to observe and quantitate the process of phagocytosis in human polynuclear cells and mouse macrophages by following the internalization of a foreign particle—in this case killed E. coli (K-12 strain) cells that have been labeled with the fluorescent dye fluorescein. In addition to the lyophilized E. coli BioParticles® component, the kit contains a trypan blue solution (to quench the fluorescence from particles that were not internalized) as well as step-by-step instructions for performing this phagocytosis assay in a fluorescence microplate reader. The methodology used with this kit has been developed using an adherent murine macrophage cell line (J774); however, by modifying the cell culture conditions, researchers can adapt this phagocytosis assay to other adherent cell lines.

Vybrant® Phagocytosis Assay Kit Specifications:
• Particles are fluorescein-labeled E. coli (K-12 strain)
• Kit supplies sufficient reagents for ~250 tests using 96-well plates
• Fluorescence is measured using a microplate reader (Ex/Em ~480/520 nm)


Find More BioParticles® Products
We offer a large range of dye-labeled and unlabeled E. coli (K-12 strain), S. aureus (Wood strain, without protein A), and zymosan (S. cerevisiae) BioParticles® products. Find out about these products and their applications by reviewing Probes for Following Receptor Binding and Phagocytosis—Section 16.1 in the Molecular Probes® Handbook.

For pH-sensitive endocytosis assays, see our pHrodo™ BioParticles® conjugates.

For Research Use Only. Not for human or animal therapeutic or diagnostic use.

eBioscience™ Essential Human Th1/Th17 Phenotyping Kit (Invitrogen™)

The Invitrogen eBioscience Essential Human Th1/Th17 Phenotyping Kit is designed for flow cytometry-based identity testing of Type 1 and Type 17 T–helper cells (Th1/Th17). It leverages high quality eBioscience antibodies, isotype controls, cell stimulation reagents (induces cytokine production), and fixation/permeabilization reagents (for intracellular markers IfN-g and IL-17) and includes a ready-to-use protocol for experimental setup, gating strategy, and analysis.

Benefits include:
Proven performance―antibodies and supporting reagents designed for flow cytometry
Highly referenced antibodies―includes one of the most referenced clones for IL-17
Turnkey kit and protocol―contains reagents and standardized gating recommendations needed to perform flow cytometry characterization

CellEvent™ Caspase-3/7 Green Detection Reagent (Invitrogen™)

CellEvent™ Caspase-3/7 Green Detection Reagent is a novel fluorogenic substrate for activated caspase-3/7 that is compatible with both live cell and fixed imaging, with absorption/emission maxima of ~502/530 nm. Activation of caspase-3 is an essential event during apoptosis, making this an optimized reagent for analysis of apoptotic cells.

See more tools for detection of apoptotic cells >

• Optimized caspase-3/7 substrate for apoptosis analysis
• Easy to use—add and read; no cell lysis required
• Use for time course measurements; easily select the time point of interest
• Compatible with both live cell fluorescence-imaging and formaldehyde-based fixation methods
• Multiplex enabled—combine with other fluorescent reagents to confirm apoptosis in the same cell or cell population

Substrate specifics
CellEvent Caspase-3/7 Green Detection Reagent is a four-amino acid peptide (DEVD) conjugated to a nucleic acid-binding dye with absorption/emission maxima of ~502/530 nm. The DEVD peptide sequence is a cleavage site for caspase-3/7, and the conjugated dye is non-fluorescent until cleaved from the peptide and bound to DNA.

Principles of action
CellEvent Caspase-3/7 Green Detection Reagent is intrinsically non-fluorescent as the DEVD peptide inhibits the ability of the dye to bind to DNA. However, after activation of caspase-3/7 in apoptotic cells, the DEVD peptide is cleaved, enabling the dye to bind to DNA and produce a bright, fluorogenic response. The fluorescent emission of the dye when bound to DNA is ~530 nm and can be observed using a standard FITC filter set.

Simple three-step protocol
To use CellEvent Caspase 3/7 Detection Reagent, simply add it to cells, incubate 30 minutes, and visualize (see figure). Apoptotic cells with activated caspase-3/7 will have bright green nuclei, while cells without activated caspase 3/7 will have minimal fluorescent signal (see figure).

Assay versatility allows for live cell detection and fixation
This robust assay enables the examination of caspase-3/7 activation in live cells. Furthermore, since wash steps are not necessary for detection, fragile apoptotic cells typically lost during wash steps are preserved. Importantly, the fluorescent signal from CellEvent Caspase 3/7 Detection Reagent can survive fixation and permeabilization. This allows for the flexibility to perform end-point assays and probe for other proteins of interest using immunocytochemistry.

eBioscience™ Essential Human T-Cell Phenotyping Kit (Invitrogen™)

The Invitrogen eBioscience Essential Human T-Cell Phenotyping Kit is designed for flow cytometry-based identity testing of T-cells. It leverages high-quality eBioscience antibodies and isotype controls and includes a ready-to-use protocol for experimental setup, gating strategy, and analysis.

Benefits include:
Proven performance―antibodies and supporting reagents designed for flow cytometry
Highly referenced antibodies―eBioscience portfolio contains trusted fluorochrome-conjugated antibodies and reagents for immunology and oncology research
Turnkey kit and protocol―contains reagents and standardized gating recommendations needed to perform flow cytometry characterization

eBioscience™ BrdU Kit for IHC/ICC Immunofluorescence eFluor™ 660 (Invitrogen™)

This BrdU Kit for immunohistochemistry (IHC)/immunocytochemistry (ICC) with immunofluorescence detection contains the necessary reagents and buffers for identifying and examining proliferating cells by immunohistochemical or immunocytochemical analysis. Cells are labeled in vitro or in vivo with 5-bromo-2’-deoxyuridine (BrdU), a synthetic analog of thymidine, which is incorporated into DNA in place of thymidine during the S-phase of the cell cycle. Following fixation and antigen retrieval steps, cells or tissue sections are stained for BrdU incorporation and visualized using a streptavidin-conjugated fluorophore. This kit has been optimized for IHC with both frozen and formalin-fixed paraffin embedded BrdU-labeled mouse intestine and ICC of BrdU-pulsed HeLa cells grown on culture slides.

Conjugate
eFluor 660

Laser
Red Laser

Emit
668 nm

Excite
633 - 647 nm

Reported Application
Immunohistochemical Staining of Formalin-Fixed Paraffin Embedded Tissue Sections, Immunohistochemical Staining of Frozen Tissue Sections, Immunocytochemistry, Microscopy

Vybrant™ Apoptosis Assay Kit #5, Hoechst 33342/Propidium Iodide (Invitrogen™)

The Vybrant Apoptosis Assay Kit #5 provides a rapid and convenient assay for apoptosis based on fluorescence detection of the compacted state of the chromatin in apoptotic cells. This kit contains ready-to-use solutions of blue-fluorescent Hoechst 33342 dye, which stains the condensed chromatin of apoptotic cells more brightly than the chromatin of nonapoptotic cells, and red-fluorescent propidium iodide dye, which stains dead cells.

Premo™ Autophagy Sensor LC3B-GFP (BacMam 2.0) (Invitrogen™)

Premo™ Autophagy Sensor combines the selectivity of a LC3B-green fluorescent protein (GFP) chimera with the transduction efficiency of BacMam technology, enabling unambiguous visualization of this protein. BacMam reagents (insect Baculovirus with a Mammalian promoter) are non-replicating in mammalian cells and thus safe to handle. They are also non-cytotoxic and ready-to-use. Unlike expression vectors, BacMam reagents enable titratable and reproducible expression and offer high co-transduction efficiency, therefore multiple BacMam reagents can readily be used in the same cell. Recent improvements made to the BacMam system enable efficient transduction in a wider variety of cells including neurons and neural stem cells (NSCs) with an easy, one-step protocol. Now, to visualize autophagy, simply add the BacMam LC3B-GFP to the cells and incubate overnight for protein expression. Each Premo™ Autophagy Sensor Kit includes the BacMam LC3B-FP, a control BacMam LC3B (G120A)-FP (mutation on the control BacMam LC3B prevents cleavage and subsequent lipidation during normal autophagy, thus protein localization should remain cytosolic and diffuse) and chloroquine diphosphate to artificially induce autophagosome accumulation.

BacLight™ RedoxSensor™ Green Vitality Kit (Invitrogen™)

The RedoxSensor™ Green reagent included in the BacLight™ RedoxSensor™ Green Vitality Kit is an indicator of bacterial reductase activity. This reductase activity is, in turn, a reliable marker for changes in electron transport chain function and for changes in vitality that occurs following antibiotic treatment. RedoxSensor™ Green reagent penetrates both gram-positive and gram-negative bacteria. Following reduction, the RedoxSensor™ Green reagent will produce a stable green-fluorescent signal in 10 minutes that is compatible with formaldehyde fixation techniques.

View additional information about all microbiology assays for flow cytometry.

eBioscience™ Mouse Th17 Cytokine Staining Panel (Invitrogen™)

This mouse Th17 cytokine staining panel includes all reagents needed for simultaneous flow cytometric detection of all the major cytokines produced by the Th17 lineage: IL-17A, IL-17F, IL-21 and IL-22. An anti-CD4 antibody that can be used after fixation and permeabilization, as well as IC Fixation and Permeabilization Buffers, is also included.

CD4+ T helper cells are critical mediators of the cellular immune response. For many years, due to cytokine expression patterns, it was thought that CD4+ T helper cells existed as a dichotomy of lineages named Th1 and Th2. However, further investigation revealed that the T helper cell population was not limited to these two subsets. Although it had long been appreciated that IL-17 (also known as IL-17A) production by T cells was required for protection against some pathogens, studies demonstrated that this cytokine was produced by a unique subset of T helper cells. Subsequent reports definitively showed that T cells could differentiate into IL-17-producing cells in vitro and in vivo independent of Th1 or Th2 development, thereby establishing Th17 cells as a unique T helper cell lineage. In addition to IL-17A expression, Th17 cells have been reported to express IL-17F (and heterodimers of Il-17 and Il-17F), IL-21 and IL-22. Functionally, Th17 cells play a role in host defense against extracellular pathogens by mediating the recruitment of neutrophils and macrophages to infected tissues. Moreover, it is becoming evident that aberrant regulation of Th17 cells may play a significant role in the pathogenesis of multiple inflammatory and autoimmune disorders.

Reactivity/Species
Mouse

Reported Application
Intracellular Staining Followed by Flow Cytometric Analysis

Escherichia coli (K-12 strain) BioParticles™, fluorescein conjugate (Invitrogen™)

The Molecular Probes® BioParticles® product line consists of a series of fluorescently labeled, heat- or chemically killed bacteria and yeast in a variety of sizes, shapes, and natural antigenicities. These fluorescent BioParticles® products have been employed to study phagocytosis by fluorescence microscopy, quantitative spectrofluorometry, and flow cytometry.

We offer E. coli (K-12 strain), S. aureus (Wood strain, without protein A) and zymosan (S. cerevisiae) BioParticles® conjugates covalently labeled with a variety of different fluorophores (special care has been taken to remove free dye after conjugation). Unlike the fluorescence of fluorescein-labeled BioParticles® conjugates, which is partially quenched in acidic environments, the fluorescence of the Alexa Fluor®, BODIPY® FL, tetramethylrhodamine and Texas Red® dye conjugates is uniformly intense over the pH range from 3 to 10.

BioParticles Specifications:
• Label (Ex/Em): Fluorescein (~494/518 nm)
• Particle: E. coli (K-12 strain)
Opsonizing reagent available


Using BioParticles Products
BioParticles® conjugates are provided as lyophilized powders. There are approximately 3 x 108 E. coli or S. aureus particles per mg solid and approximately 2 x 107 zymosan particles per mg solid. BioParticles® conjugates can be reconstituted in the buffer of your choice for use in phagocytosis assays. The fluorescence of BioParticles® conjugates that are bound to the surface of the cell (but not internalized) can be quenched by ethidium bromide, trypan blue, or other quenchers. In addition to cellular applications, fluorescent BioParticles® conjugates may be effective as flow cytometry calibration references when sorting bacteria and yeast mutants. These small particles may also be useful references for light scattering studies because their sizes and shapes differ in characteristic ways.

Find More BioParticles® Products
We offer a large range of dye-labeled and unlabeled E. coli (K-12 strain), S. aureus (Wood strain, without protein A), and zymosan (S. cerevisiae) BioParticles® products. Find out about these products and their applications by reviewing Probes for Following Receptor Binding and Phagocytosis—Section 16.1 in the Molecular Probes® Handbook.

For pH-sensitive endocytosis assays, see our pHrodo™ BioParticles® conjugates.

For Research Use Only. Not for human or animal therapeutic or diagnostic use.

Arcturus™ Paradise™ PLUS 2 Round Kit (Applied Biosystems™)

The Arcturus® Paradise® PLUS Reagent System permits unprecedented gene expression analysis of millions of previously inaccessible samples. It is an integrated solution for gene expression studies from formalin-fixed, paraffin-embedded (FFPE) tissues, including reagents for tissue staining, RNA isolation and RNA amplification. Amplify RNA from as little as 5 ng of fixed RNA using methods that have been specially optimized to overcome the challenges of cross-linked templates.

This kit provides reagents for 2-round in-vitro transcription.

Key product features:
• Accelerated discovery process – unlock the tissue archives to access previously inaccessible samples
• Simplified procedure – achieve unprecedented results in six steps
• Streamlined application – integrate seamlessly into current workflow
• Efficient workflow – secure downstream success with the Paradise® QC Kit
• Comprehensive system – benefit from the only complete system for formalin-fixed samples

Unlock the Tissue Archives
Currently, tens of millions of tissue samples with known outcomes are available in tissue archives. The Paradise® PLUS Reagent System enables microarray and qRT-PCR analysis of these previously inaccessible samples, allowing retrospective studies to accelerate discoveries.

Achieve Unprecedented Results in Six Steps
Only six steps are required to take you from sample to results (See Figure 1).

Integrate Seamlessly into Current Workflow
The Paradise® PLUS Reagent System is compatible with all major microarray platforms. Robust and reproducible results are achieved using tools and techniques commonly employed in today’s research laboratories (See Figures 2, 3, 4, and 5).

Secure Downstream Success with the Paradise® QC Kit
Not all FFPE samples contain high quality RNA. Identify the best samples for your studies using a simple, real-time PCR research technique, and then proceed with confidence using these pre-qualified samples (See Figure 6).

Benefit from the Only Complete System for Formalin-fixed Samples
The Paradise® PLUS Reagent System is the only available solution that provides all the components necessary to prepare RNA from formalin-fixed, paraffin-embedded tissue samples for gene expression analysis. To ensure successful microarray results, the system includes reagents optimized for exceptional recovery of RNA and superior RNA amplification.

Integrated Systems for Microgenomics
The Paradise PLUS Reagent System is a validated as part of the Arcturus® Complete System for Microgenomics®, an integrated solution for utilizing small quantities of RNA for gene expression analysis. This system features the ArcturusXT™ Laser Capture Microdissection (LCM) Instrument to procure pure cell populations. Arcturus Systems for Microgenomics enable accurate and sensitive microarray assays that reveal molecular signatures otherwise obscured in whole tissue samples or cell mixtures.

For Research Use Only. Not for use in diagnostics procedures.

Vybrant™ FAM Caspase-8 Assay Kit, for flow cytometry (Invitrogen™)

The Vybrant™ FAM Caspase-8 Assay Kit employs a novel approach to detect active caspases that is based on a fluorescent inhibitor of caspases (FLICA™) methodology, essentially an affinity label. The reagent associates a fluoromethyl ketone (FMK) moiety, which can react covalently with a cysteine, with a caspase-specific amine acid sequence. For caspase-8, this recognition sequence is leucine-glutamic acid-threonine-aspartic acid (LETD). A fluorescein group is attached as a reporter. The FLICA reagent is thought to interact with the enzymatic reactive center of an activated caspase via the recognition sequence, and then to attach covalently through the FMK moiety. The FLICA inhibitor is cell permeant and noncytotoxic. Unbound FLICA molecules diffuse out of the cell and are washed away; the remaining green-fluorescent signal is a direct measure of the amount of active caspase that was present at the time the inhibitor was added. This kit includes the FLICA reagent specific for caspase-8, Hoechst 33342 stain, and propidium iodide stain, which allows the simultaneous evaluation of caspase activation, membrane permeability, and cell cycle by flow cytometry.

View a selection guide for all apoptosis assays for flow cytometry.

Amplex™ Red Xanthine/Xanthine Oxidase Assay Kit (Invitrogen™)

The Amplex Red Xanthine/Xanthine Oxidase Assay Kit provides a sensitive and simple fluorometric method for detecting as little as 200 nM xanthine or hypoxanthine or as little as 0.1 mU/mL of xanthine oxidase activity in a purified system with a 100 µL assay volume.

Escherichia coli (K-12 strain) BioParticles™, Alexa Fluor™ 488 conjugate (Invitrogen™)

The Molecular Probes® BioParticles® product line consists of a series of fluorescently labeled, heat- or chemically killed bacteria and yeast in a variety of sizes, shapes, and natural antigenicities. These fluorescent BioParticles® products have been employed to study phagocytosis by fluorescence microscopy, quantitative spectrofluorometry, and flow cytometry.

We offer E. coli (K-12 strain), S. aureus (Wood strain, without protein A) and zymosan (S. cerevisiae) BioParticles® conjugates covalently labeled with a variety of different fluorophores (special care has been taken to remove free dye after conjugation). Unlike the fluorescence of fluorescein-labeled BioParticles® conjugates, which is partially quenched in acidic environments, the fluorescence of the Alexa Fluor®, BODIPY® FL, tetramethylrhodamine and Texas Red® dye conjugates is uniformly intense over the pH range from 3 to 10.

BioParticles Specifications:
• Label (Ex/Em): Alexa Fluor® 488 (~495/519 nm)
• Particle: E. coli (K-12 strain)
Opsonizing reagent available


Using BioParticles Products
BioParticles® conjugates are provided as lyophilized powders. There are approximately 3 x 108 E. coli or S. aureus particles per mg solid and approximately 2 x 107 zymosan particles per mg solid. BioParticles® conjugates can be reconstituted in the buffer of your choice for use in phagocytosis assays. The fluorescence of BioParticles® conjugates that are bound to the surface of the cell (but not internalized) can be quenched by ethidium bromide, trypan blue, or other quenchers. In addition to cellular applications, fluorescent BioParticles® conjugates may be effective as flow cytometry calibration references when sorting bacteria and yeast mutants. These small particles may also be useful references for light scattering studies because their sizes and shapes differ in characteristic ways.

Find More BioParticles® Products
We offer a large range of dye-labeled and unlabeled E. coli (K-12 strain), S. aureus (Wood strain, without protein A), and zymosan (S. cerevisiae) BioParticles® products. Find out about these products and their applications by reviewing Probes for Following Receptor Binding and Phagocytosis—Section 16.1 in the Molecular Probes® Handbook.

For pH-sensitive endocytosis assays, see our pHrodo™ BioParticles® conjugates.

For Research Use Only. Not for human or animal therapeutic or diagnostic use.

Click-iT™ Plus EdU Pacific Blue™ Flow Cytometry Assay Kit (Invitrogen™)

The Click-iT® Plus EdU Pacific Blue™ Flow Cytometry Assay Kit provides a simplified, more robust assay for analyzing DNA replication in proliferating cells compared to traditional BrdU methods. Newly synthesized DNA is analyzed using the 405 nm laser of the flow cytometer. The Click-iT® Plus formulation is compatible with standard fluorophores, including R-PE and R-PE tandems, as well as fluorescent proteins.

• Multiplexable—compatible with R-PE (and tandems) and fluorescent proteins
• Accurate—superior results compared to BrdU assays
• Fast—results in as little as 60 minutes

View selection guide for all Click-iT™ EdU and Click-iT™ Plus EdU assays for flow cytometry.

Multiplexable
The Click-iT® Plus formulation provides increased multiplexibility compared to the original Click-iT® EdU Flow Cytometry assays. Click-iT® Plus EdU assays can be used in conjunction with R-PE and R-PE tandems, as well as fluorescent proteins such as GFP and mCherry, without loss of the accuracy or speed of the original Click-iT® EdU assay.

An Advanced Method Giving You Results Superior to BrdU
The most accurate method of proliferation analysis is direct measurement of DNA synthesis. Originally, this was performed through incorporation of radioactive nucleosides. This method was replaced by antibody-based detection of the nucleoside analog bromodeoxyuridine (BrdU). The Click-iT® Plus EdU Flow Cytometry assay is a novel alternative to the BrdU assay. EdU (5-ethynyl-2´-deoxyuridine) is a thymidine analog that is incorporated into DNA during active DNA synthesis. Detection is based on click chemistry, which is a copper-catalyzed covalent reaction between an azide and an alkyne. In this application, the alkyne is found in the ethynyl moiety of EdU, while the azide is coupled to the Alexa Fluor® dye. Standard flow cytometry methods are used for determining the percentage of S-phase cells in the population.

Mild Conditions Allow Use with Cell Cycle Dyes and Antibodies
The small size of the dye azide allows for efficient detection of the incorporated EdU using mild conditions, while standard aldehyde-based fixation and detergent permeabilization are sufficient for the Click-iT® Plus detection reagent to gain access to the DNA. This is in contrast to BrdU assays that require DNA denaturation (using HCl, heat, or digestion with DNase) to expose the BrdU so that it can be detected with an anti-BrdU antibody. Sample processing for the BrdU assay can result in signal alteration of the cell cycle distribution, as well as destruction of antigen recognition sites when using the HCl method. In contrast, the easy-to-use Click-iT® Plus EdU assay is compatible with cell cycle dyes. The Click-iT® Plus EdU assay can also be multiplexed with antibodies against surface and intracellular markers, as well as conjugates labeled with standard fluorophores including R-PE, R-PE tandems, and fluorescent proteins (GFP and mCherry).

Quick and Simple Protocol
The Click-iT® Plus EdU protocol is based on the aldehyde fixation and detergent permeabilization steps for immunohistochemical antibody labeling. However, EdU is compatible with other fixation/permeabilization agents including saponin and methanol. In just five steps you’ll be ready to analyze your cell proliferation data:
1. Treat cells with EdU.
2. Fix and permeabilize cells.
3. Detect S-phase cells with Click-iT® Plus detection cocktail for 30 min.
4. Wash once.
5. Analyze.
Results can be seen in as little as 60 minutes in some circumstances, but we recommend 90 minutes for all applications.

HCS DNA Damage Kit (Invitrogen™)

The HCS DNA Damage kit simultaneously and quantitatively measures two important cell-health parameters: DNA damage and cytotoxicity. DNA damage is detected using an antibody against phosphorylated H2AX (Ser139) which is induced in response to double-strand break (DSB) formation. To detect cytotoxicity, the Image-iT® DEAD™ Green is an impermeant dye in healthy cells that becomes permeant when the plasma membrane of cells is compromised. Both indicators are amenable to fixation and permeabilization, thus allowing for multiplexing with other biomarkers of interest. The blue-fluorescent nuclear segmentation tool Hoechst 33342 is also included and stains DNA in both live and dead cells.

eBioscience™ Human/Non-Human Primate Regulatory T Cell Staining Kit #1 (Invitrogen™)

This Human/Non-human Primate Regulatory T cell Staining Kit contains all of the buffers and monoclonal antibodies for CD4, CD25, and Foxp3 necessary to successfully stain and identify regulatory T cells from human, rhesus, cynomolgus, or chimpanzee peripheral blood cells. This kit may also be used to identify regulatory T cells in baboons, using the CD25 and Foxp3 monoclonal antibodies, however, the monoclonal antibody for CD4 that is included in this kit does not crossreact with baboon.

The OKT4 monoclonal antibody reacts with human, rhesus, cynomolgus, and chimpanzee CD4, a 59 kDa glycoprotein found on the surface of the majority of thymocytes, a subset of mature T cells (T helper cells), and at lower levels on monocytes. The BC96 monoclonal antibody reacts with human, rhesus, cynomolgus, chimpanzee, and baboon CD25 (also known as interleukin-2 receptor alpha, IL-2R alpha), a 55 kDa surface protein expressed by early progenitors of T cells and B cells, by mature, activated T cells and B cells, and at constitutively high levels on regulatory T cells. The PCH101 monoclonal antibody reacts with the amino terminus of human, rhesus, cynomolgus, chimpanzee, and baboon Foxp3 also known as FORKHEAD BOX P3, SCURFIN, and JM2. Foxp3 is a 49-55 kDa protein and a member of the forkhead/winged-helix family of transcription factors. It was identified as the gene responsible for the X-linked lymphoproliferative disease observed in scurfy (sf) mice and in the human disorder, X-linked autoimmunity-allergic dysregulation syndrome (XLAAD). Constitutive expression of Foxp3 mRNA has been shown in CD4+CD25+ regulatory T cells (Treg), and ectopic expression of Foxp3 in CD4+CD25- cells imparts a Treg phenotype in these cells.

Reactivity/Species
Human, Monkey

Reported Application
Intracellular Staining Followed by Flow Cytometric Analysis

Pierce™ Peroxidase IHC Detection Kit (Thermo Scientific™)

The Thermo Scientific Pierce Peroxidase IHC Detection Kit bundles the Pierce Metal Enhanced DAB Substrate Kit with all necessary components to stain, counterstain and preserve experimental results with frozen or paraffin tissue sections.

Features of the Peroxidase IHC Detection Kit:

Incredibly sensitive—fifty times more sensitive than the traditional DAB (3,3'-diaminobenzidine) method and 30 times more sensitive than other metal-intensified versions of DAB
Low background, high intensity, large dynamic range—gives a crisp dark brown-black precipitate that's more intense than the dull brown precipitate observed when using DAB without enhancement; substrate delivers increased intensity with almost nonexistent background
Easy-to-use, two-component system—mix the two liquid components and the working solution is ready-to-use; sufficient substrate and other staining reagents to treat up to 500 slides
Six-hour working solution stability—unlike other DAB substrates that must be used immediately, the working solution is stable for more than six hours at room temperature
Complete kit—saves time in reagent ordering, gathering and validating reagents for the IHC application; saves money compared to purchasing of all components separately

This immunohistochemical staining kit bundles the Pierce Metal Enhanced DAB Substrate Kit with all necessary components to stain, counter-stain and preserve experimental results with frozen or paraffin tissue section that have been probed with peroxidase enzyme conjugates. The Pierce Peroxidase IHC Detection Kit eliminates the need to procure numerous reagents from various sources, resulting in a time- and cost-effective way to perform peroxidase-based tissue staining. Harris Hematoxylin nuclear stain and mounting medium allow counter-staining and tissue preservation, respectively. The detection method is compatible with direct or avidin-biotin-complex (ABC) probing strategies.

Applications:
• Tissue immunohistochemistry procedures designed for DAB precipitating substrate
• DAB staining of fixed paraffin sections, smears and frozen section preparations
• Immunohisto- or immunocyto-staining using the metal-enhanced DAB substrate

Pacific Blue™ Annexin V/SYTOX™ AADvanced™ Apoptosis Kit, for flow cytometry (Invitrogen™)

This product detects the externalization of phosphatidylserine in apoptotic cells using recombinant annexin V conjugated to violet-fluorescent Pacific Blue™ dye and dead cells using SYTOX™ AADvanced™ stain. After staining a cell population with Pacific Blue™, annexin V and SYTOX™ AADvanced™, apoptotic cells show violet fluorescence, dead cells show red fluorescence, and live cells show little or no fluorescence. These populations are easily distinguished by a flow cytometer with the 405 nm and 488 nm lines for excitation. There is very little spectral overlap between the two dyes, therefore very little compensation is needed. Each kit contains sufficient reagents for approximately 50 flow cytometry tests.

View a selection guide for all apoptosis assays for flow cytometry.

Tali™ Cell Cycle Kit (Invitrogen™)

The Tali® Cell Cycle Kit includes a ready-to-use reagent that, when used with the Tali® Image-Based Cytometer, provides a convenient and accurate determination of the percentage of cells in each phase of the cell cycle. This single-addition reagent is composed of propidium iodide (PI), RNase A, and Triton X-100.

The Tali® Cell Cycle Kit offers a simple workflow:

• The pre-mixed reagent can be stored at room temperature and used when your cells are ready
• Simply fix your cells, add the reagent, incubate, and visualize
• Cell cycle data can be analyzed on the Tali® instrument or exported as a .csv or .fcs file for analysis with various modeling software

Fluorescent detection of cellular DNA as a cell-cycle phase indicator
PI, a red fluorescent dye, binds to DNA by intercalating between the bases with the little or no sequence preference. Consequently, the degree of fluorescence is proportional to the amount of cellular DNA, which itself is indicative of cell cycle phase (as cells progress through the cell cycle, the amount of DNA ultimately doubles). PI will also label RNA, so the addition of RNase A is necessary for accurate determination of the percentage of cells in each phase of the cell cycle.

After fixing your cells, simply add the optimized Tali® Cell Cycle reagent, incubate for 30 minutes in the dark, and then analyze on the Tali® Image-Based Cytometer. The cell cycle data from the Tali® Image-Based Cytometer can be analyzed on the instrument or exported in either .csv or .fcs format, and analyzed using cell cycle modeling software (such as ModFit or MultiCycle).

Amplex™ Red Sphingomyelinase Assay Kit (Invitrogen™)

The Amplex® Red Sphingomyelinase Assay Kit provides a sensitive, rapid, and simple fluorometric method for detecting very low concentrations of sphingomyelinase using a fluorescence microplate reader or fluorometer.

See our complete line of Fluorescence Microplate assays.

• Detects sphingomyelinase activity levels as low as 80 µU/mL in a 200 µL assay volume
• Format allows for multiple time point measurements
• Designed for minimal autofluorescence interference

In this enzyme-coupled assay, sphingomyelinase activity is monitored indirectly using 10-acetyl-3,7-dihydroxyphenoxazine (Amplex® Red reagent), a sensitive fluorogenic probe for hydrogen peroxide. Sphingomyelinase hydrolyses the sphingomyelin to yield ceramide and phosphorylcholine. Alkaline phosphatase is added, which hydrolyzes phosphorylcholine to form choline. The choline is then oxidized by choline oxidase to form betaine and hydrogen peroxide. In the presence of horseradish peroxidase, the hydrogen peroxide reacts with Amplex® Red reagent in a 1:1 stoichiometric ratio to generate the highly fluorescent product resorufin.

Because resorufin has absorption and fluorescence emission maxima of approximately 571 nm and 585 nm, respectively, there is little interference from autofluorescence in most biological samples.

Use Amplex® Red Assays for a Broad Range of Investigations
A wide variety of validated Amplex® Red assays are available for studying cell signaling and lipids, neurobiology, inflammation and immune function, and metabolism. We also offer Amplex® UltraRed Reagent (Cat. No. A36006), a second-generation reagent providing greater sensitivity and brighter fluorescence, and the Amplex® Red/UltraRed Stop Reagent (Cat. No. A33855). The Amplex® Red/UltraRed Stop Reagent provides convenience and control by allowing the fluorescence signal-generating reaction to be terminated at a user-determined time point. After addition of the stop reagent, the fluorescence signal remains stable for at least three hours. Custom assay design and packaging are also available.

ThiolTracker™ Violet (glutathione detection reagent) (Invitrogen™)

ThiolTracker™ Violet is a brighter and more robust intracellular thiol probe for reproducible GSH detection. Since reduced glutathione represents the majority of intracellular free thiols in the cell, ThiolTracker™ Violet can be used in estimating the cellular level of reduced glutathione.
ThiolTracker™ Violet offers the following;
• 10X brighter than mBCL
• UV or 405 nm excitation, 525 nm emission
• Easy to use
• Reproducible results
• Can be fixed with aldehydes and permeabilized by Triton® X-100 (0.5%)
• Formaldehyde fixable and suvives detergent extraction
• Can be used in multiplex assays including cytotoxicity studies
Subcellular detection and localization of GSH is important in understanding the modulation of redox status, the effect of drugs, and the mechanisms of detoxification. Furthermore, differences in GSH levels in response to oxidative stress in subpopulations of cells have been reported, underscoring the importance of detection methods amenable to flow cytometry and automated fluorescence detection. ThiolTracker™ Violet can be used for Flow Cytometry, HCS imaging, and epifluorescent microscopy.

Histogene™ Refill Kit, includes dehydration chemicals & stain only (Applied Biosystems™)

This kit contains only the dehydration chemicals and stain used with the ARCTURUS® HistoGene® LCM Frozen Section Staining Kit. Designed as a fast-penetrating stain, this kit provides excellent contrast while preserving nucleic acid integrity and retaining low-abundance mRNAs.

Key product features:
• Optimized process—provide superior staining while preserving RNA
• Comprehensive kit—simplify tissue staining and dehydration
• High-quality RNA yield—retain low-abundance mRNA and maintain RNA integrity
• Versatile application—obtain high-quality RNA from many tissue types
• Modular design—use with other ARCTURUS® products to produce superior microarray data

Provide Superior Staining while Preserving Nucleic Acids
HistoGene® Staining Solution has been developed by ARCTURUS® to stain tissues for LCM subsequently used as sources of nucleic acids. It is a fast-penetrating stain that provides good contrast by differential staining of nuclei (purple) and cytoplasm (light pink). By minimizing the exposure of tissues to water where nucleases may be activated, HistoGene® stain helps to preserve nucleic integrity that may be otherwise compromised when using longer staining protocols.

Retain Low-abundance mRNA
RT-PCR analysis of specific genes from cells captured from tissues processed using the HistoGene® LCM Frozen Section Staining Kit shows retention of both low-abundance mRNA and higher-abundance species. The mRNA profile of HistoGene® samples appears free of degradation.

Maintain RNA Integrity
Tissues prepared for LCM using HistoGene® LCM Frozen Section Staining Kit reagents, supplies and instructions yield high quality RNA.

Obtain High-quality RNA from Many Tissue Types
ARCTURUS® has validated the HistoGene® Kit by examining RNA integrity on many tissue types. All tissues tested to date using the HistoGene® Kit have yielded quality RNA. Validated tissue types include (Table 1):

Part of the Arcturus® System for Microgenomics
The HistoGene® LCM Frozen Section Staining Kit is part of the ARCTURUS® Complete System of Microgenomics and is designed to seamlessly work together to produce high quality expression microarrays. With the PicoPure® RNA Isolation Kit, you can recover a high yield of RNA from as few as 10 cells in a minimal volume. The RiboAmp® RNA Amplification Kit can amplify picograms of your RNA sample into micrograms of amplified RNA (aRNA) that is ready for labeling and hybridization to microarrays. ARCTURUS® instruments and kits increase the reliability and reproducibility of gene expression studies.

For Research Use Only. Not for use in diagnostics procedures.

eBioscience™ Human Regulatory T Cell Staining Kit (Invitrogen™)

This Human Regulatory T cell Staining Kit contains all of the buffers and monoclonal antibodies for CD4, CD25, and Foxp3 necessary to successfully stain and identify regulatory T cells from human peripheral blood cells.

The RPA-T4 monoclonal antibody reacts with human CD4, a 59 kDa glycoprotein found on the surface of the majority of thymocytes, a subset of mature T cells (T helper cells), and at lower levels on monocytes. The BC96 monoclonal antibody reacts with human CD25 (also known as interleukin-2 receptor alpha, IL-2R alpha), a 55 kDa surface protein expressed by early progenitors of T cells and B cells, by mature, activated T cells and B cells, and at constitutively high levels on regulatory T cells. The PCH101 monoclonal antibody reacts with the amino terminus of human Foxp3 also known as FORKHEAD BOX P3, SCURFIN, and JM2. Foxp3 is a 49-55 kDa protein and a member of the forkhead/winged-helix family of transcription factors. It was identified as the gene responsible for the X-linked lymphoproliferative disease observed in scurfy (sf) mice and in the human disorder, X-linked autoimmunity-allergic dysregulation syndrome (XLAAD). Constitutive expression of Foxp3 mRNA has been shown in CD4+CD25+ regulatory T cells (Treg), and ectopic expression of Foxp3 in CD4+CD25- cells imparts a Treg phenotype in these cells.

Reactivity/Species
Human

Reported Application
Intracellular Staining Followed by Flow Cytometric Analysis

Click-iT™ TUNEL Alexa Fluor™ 647 Imaging Assay, for microscopy & HCS (Invitrogen™)

The Click-iT® TUNEL Alexa Fluor® 647 Imaging Assay utilizes a dUTP modified with an alkyne, a small, bio-orthogonal functional group that enables the nucleotide to be more readily incorporated by TdT than other modified nucleotides, including BrdUTP, biotin-dUTP or fluorescein-dUTP. Detection is based on a click reaction, a copper (I) catalyzed reaction between an azide and alkyne. Click chemistry fills the void when methods such as direct labeling or the use of antibodies are not efficient. The small size of the detection reagent, an Alexa Fluor® azide (MW <~1000) compared to that of an antibody (MW ~150,000) enables facile penetration of complex samples with only mild fixation and permeabilization required.

Compared to other assays using one of the other modified nucleotides, the Click-iT® TUNEL assay is able to detect a higher percentage of apoptotic cells under identical conditions. The assay is also fast and is complete within 2 hours. Furthermore, the Click-iT® TUNEL assay allows multiplexing with surface and intracellular biomarker detection.

More tools for apoptosis detection and measurement >

Ultra-Sensitive ABC Peroxidase Standard Staining Kit (Thermo Scientific™)

Thermo Scientific Pierce ABC Staining Kits include reagents for the avidin-biotin complex (ABC) technique, a highly sensitive method for immunohistochemical detection with biotinylated secondary antibodies.

Pierce ABC Staining Kits are highly sensitive, produce very low background staining and have rapid avidin-biotin interactions. Standard Kits contain only the two essential reagents (avidin and biotinylated HRP) can be used with nearly any species of biotinylated secondary antibody. The Ultra-Sensitive ABC Peroxidase Staining Kits are approximately five times more sensitive than the regular-senstivity kit, enabling more dilute primary antibodies to be used while producing equal staining intensity. Ultra-Sensitive Kits are offered as a Standard Kit and as Complete Kits, which contain biotinylated secondary antibodies and appropriate block serum for use with mouse or rabbit antibodies, respectively.

Related Products
ABC Peroxidase Standard Staining Kit
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Ultra-Sensitive ABC Peroxidase Rabbit IgG Staining Kit

Arcturus™ Paradise™ Plus staining components (Applied Biosystems™)

The ARCTURUS® Paradise® PLUS FFPE LCM Staining Kit offers a simple solution developed to stain formalin-fixed paraffin-embedded (FFPE) tissues before proceeding with laser capture microdissection (LCM) and downstream molecular analysis. FFPE samples introduce unique challenges for gene expression profiling, including chemical modification and fragmentation of RNA molecules. Archival of FFPE samples adds to these challenges through increased RNA degradation over time.

Key product features:
• Provide Superior Staining while Preserving RNA
• Secure Downstream Success with the Paradise® QC Kit
• Benefit from the Only Complete System for Formalin-fixed Samples

Provide Superior Staining while Preserving RNA
The Paradise® PLUS FFPE LCM Staining Kit offers a simple solution developed to stain FFPE tissues before proceeding with laser microdissection and downstream molecular analysis. It is a fast penetrating stain that provides good contrast by differential staining of nuclei (purple) and cytoplasm (light pink). By minimizing the exposure of tissues to water where nucleases may be activated, the Paradise® PLUS FFPE LCM Staining Kit helps to preserve RNA integrity that may be otherwise compromised when using longer staining protocols.

Secure Downstream Success with the Paradise® QC Kit
Identify the best samples for your studies using the Paradise® PLUS Sample Quality Assessment Kit (QC Kit). The QC Kit uses a simple, real-time PCR technique to check the quality of RNA in your FFPE sample before staining and LCM. Proceed with confidence knowing your FFPE samples have high quality RNA.

Benefit from the Only Complete System for Formalin-fixed Samples
The Paradise® PLUS Reagent System is the only available solution that provides all the components necessary to prepare RNA from FFPE tissue samples for gene expression analysis. To ensure successful microarray results, the system includes reagents optimized for exceptional recovery of RNA and superior RNA amplification.

Integrated Systems for Microgenomics
The Paradise® PLUS Reagent System is validated as part of the ARCTURUS® System for Microgenomics, an integrated solution for utilizing small quantities of RNA for gene expression analysis. ARCTURUS® Systems for Microgenomics enable accurate and sensitive microarray assays that reveal molecular details otherwise obscured in whole tissue samples or cell mixtures.

For Research Use Only. Not for use in diagnostics procedures.

pHrodo™ Green Zymosan Bioparticles™ Conjugate for Phagocytosis (Invitrogen™)

New pH-sensitive pHrodo® Green dye conjugates, like our pHrodo® Red dye conjugates, give faster and more accurate results than any other phagocytosis assay. pHrodo® Green conjugates are non-fluorescent outside the cell at neutral pH, but fluoresce brightly green at acidic pH such as in phagosomes. Get faster staining and more accurate results—without the need for wash steps or quencher dye.

• Specific detection of phagocytosis and endocytosis
• Reduced signal variability and improved timing in sensitive experiments
• Enough to do 100 tests when using 100 µL in each well of a 96-well plate
• Multiplex with other compatible dyes, such as red florescent protein, TMRM, NucBlue™ Hoechst, and CellROX® Deep Red

The fluorescence of the novel pHrodo® Green dye dramatically increases as pH decreases from neutral to acidic, making it an ideal tool to study phagocytosis and its regulation by drugs and/or environmental factors. The lack of fluorescence outside the cell eliminates the need for wash steps and quencher dyes.

Use the ready-made pHrodo® Green Zymosan BioParticles® Conjugate in imaging, high content screening, high throughput screening, and flow applications. pHrodo® Green dye is also available as a conjugate of E. coli BioParticles®, S. aureus BioParticles®, or dextran 10,000. To create other conjugates, such as antibody conjugates, use pHrodo® Green STP ester or pHrodo® Green maleimide.

For Research Use Only. Not for human or animal therapeutic or diagnostic use.

CaspGLOW™ Fluorescein Active Caspase Staining Kit (Invitrogen™)

The CaspGLOW™ Fluorescein Active Staining Kit contains the reagents to sensitively detect active caspases in mammalian cells. This assay utilizes the inhibitor specific for all caspases, Z-VAD-FMK, that is directly conjugated to FITC (fluorescein isothiocyanate) for the detection system. This reagent is cell permeable, non-toxic and irreversibly binds to the active enzyme. Detection of the labeled cells can be determined by flow cytometry, fluorescent microscopy or a fluorescent plate reader.

The Caspases are a family of aspartate-specific cysteine proteases that act in a step-wise signaling manner like kinases. Recruitment of these proteases to oligomerized receptors leads to activation and autoproteolytic cleavage. Active caspases can proteolyze additional caspases generating a caspase cascade. The final outcome of this signaling pathway is a form of controlled cell death, termed apoptosis.

Reported Application
Flow Cytometric Analysis, Immunocytochemistry

Chromatin Condensation & Membrane Permeability Dead Cell Apoptosis Kit with Hoechst 33342, YO-PRO™-1, and PI dyes, for flow cytometry (Invitrogen™)

This product detects apoptotic cells with changes in nuclear chromatin condensation and plasma membrane permeability, using three nucleic acid stains: UV-excitable Hoechst 33342, green fluorescent YO-PRO™ dye, and propidium iodide. The YO-PRO™ dye can enter apoptotic cells, whereas red-fluorescent propidium iodide (PI) cannot. Thus after staining with YO-PRO™-1 dye and PI, apoptotic cells show green fluorescence, and dead cells show primarily red fluorescence and some green fluorescence. Blue fluorescent Hoechst 33342 brightly stains the condensed chromatin of apoptotic cells and more dimly stains the normal chromatin of live cells. The staining pattern resulting from the simultaneous use of these three dyes makes it possible to distinguish normal, apoptotic and dead cell populations by flow cytometry or fluorescence microscopy.

View a selection guide for all apoptosis assays for flow cytometry.

MDR1 Vesicular Transport Assay Reagent Set (Gibco™)

The MDR1 Vesicular Transport Assay Reagent Set is a ready-to-use kit containing all reagents needed for vesicular transport assay except for the vesicles themselves. The reagent set can be used for vesicular transport assay using radioisotope-labeled or non-labeled compound. This kit has enough material to perform 100 vesicular transport assays.

Use The MDR1 Vesicular Transport Assay Reagent Set to:

• Investigate the transporter interactions of your drug candidates
• Assess potential for transporter-mediated drug-drug interactions
• Obtain high quality results with a large signal to noise ratios

Concept Behind ABC Transporter Vesicles
ABC transporter vesicles are easy-to-use, efficient reagents for early assessment of a drug candidate’s substrate and drug interaction potential. While ABC transporters typically mediate the export of substrates out of cells, transporters expressed on these inside-out vesicles import substrates into the vesicles. It is therefore possible to quantitatively evaluate transport activity for your compound by determining the amount incorporated into the vesicles.

Clear and Reliable Results
Prepared from Sf9 cells which have been engineered to over-express specific ABC transporters, these "inside-out" vesicles provide high levels of transporter activity with low background, giving you a clear signal if your compound is a substrate or inhibitor of a specific efflux transporter.

Kit Compatibility
Our MDR1 Vesicular Transport Assay Reagent Set is designed for use with MDR1 Vesicles. The 100 vesicular transport assays that can be performed with this kit will require two MDR1 Vesicle products.

For Research Use Only. Not intended for animal or human therapeutic or diagnostic use.

Amplex™ Red Galactose/Galactose Oxidase Assay Kit (Invitrogen™)

The Amplex® Red Galactose/Galactose Oxidase Assay Kit provides a sensitive and simple method for detecting galactose oxidase activity or galactose using a fluorescence microplate reader or fluorometer.

See our complete line of Fluorescence Microplate assays.

• Detect concentrations as low as 2 mU/mL of galactose oxidase activity or 4 µM galactose
• Format allows for multiple time point measurements
• Designed for minimal autofluorescence interference

Galactose oxidase is an important tool in the detection of galactose in biological fluids. The Amplex® Red Galactose/Galactose Oxidase Assay Kit provides an ultrasensitive method for detecting galactose or for monitoring galactose oxidase activity. Galactose oxidase catalyzes the oxidation of galactose at the C6 position and generates hydrogen peroxide. In the presence of horseradish peroxidase (HRP), hydrogen peroxide reacts in a 1:1 stoichiometric ratio with the Amplex® Red reagent to generate the red-fluorescent compound resorufin.

Because resorufin has absorption and fluorescence emission maxima of approximately 571 nm and 585 nm, respectively, there is little interference from autofluorescence in most biological samples.

Use Amplex® Red Assays for a Broad Range of Investigations
A wide variety of validated Amplex® Red assays are available for studying cell signaling and lipids, neurobiology, inflammation and immune function, and metabolism. We also offer Amplex® UltraRed Reagent (Cat. No. A36006), a second-generation reagent providing greater sensitivity and brighter fluorescence, and the Amplex® Red/UltraRed Stop Reagent (Cat. No. A33855). The Amplex® Red/UltraRed Stop Reagent provides convenience and control by allowing the fluorescence signal-generating reaction to be terminated at a user-determined time point. After addition of the stop reagent, the fluorescence signal remains stable for at least three hours. Custom assay design and packaging are also available.

LIVE/DEAD™ Sperm Viability Kit (Invitrogen™)

The LIVE/DEAD Sperm Viability Kit provides a novel fluorescence-based assay for analyzing the viability and fertilizing potential of sperm by fluorescence microscopy or flow cytometry. Membrane-permeant SYBR® 14 nucleic acid stain labels live sperm with green fluorescence, and membrane-impermeant propidium iodide labels the nucleic acids of membrane-compromised sperm with red fluorescence.

Hypoxia Green Reagent for Flow Cytometry

Hypoxia Green Reagent for Flow Cytometry is an antibody-independent reagent used to detect low levels of oxygen levels in live cells. The membrane-permeant probe releases rhodamine as oxygen levels decrease, resulting in a fluorogenic response. Hypoxia is a characteristic of many diseases, including cardiovascular disease and tumor-mediated immunosuppression and is critical to tumor survival and growth.

• Sensitive, fixed, end-point assay
• Detects decreases in oxygen in live cells
• Fluorogenic, non-antibody based probe

Premo™ Autophagy Tandem Sensor RFP-GFP-LC3B Kit (Invitrogen™)

The Premo™ Autophagy Tandem Sensor RFP-GFP-LC3B Kit combines the ability to monitor the various stages of autophagy (through LC3B protein localization) with the high transduction efficiency and minimal toxicity of BacMam 2.0 expression technology. To perform image-based analysis for autophagy, simply add the BacMam 2.0 RFP-GFP-LC3B reagent to your mammalian cells, incubate overnight to ensure maximum protein expression, and then visualize using standard GFP (green fluorescent protein) and RFP (red fluorescent protein) settings.

This tandem RFP-GFP sensor capitalizes on the pH difference between the acidic autolysosome and the neutral autophagosome and the pH sensitivity differences exhibited by GFP (green fluorescent protein) and RFP (red fluorescent protein) to monitor progression from the autophagosome to autolysosome. The RFP and the GFP genes included in this chimera are TagRFP and Emerald GFP, respectively.

Each Premo™ Autophagy Tandem Sensor RFP-GFP-LC3B Kit includes BacMam 2.0 RFP-GFP-LC3B reagent and chloroquine diphosphate, which is used to inhibit autophagy.

Multiplex inspired: combining the Premo™ Autophagy Tandem Sensor with the far-red emitting LysoTracker® Deep Red (available separately) allows for a three-color analysis of the complete autophagic pathway dynamics
Highly efficient delivery system: >90% transduction of a wide range of mammalian cell lines, including primary cells, stem cells, and neurons
Fast and convenient: simply add the Premo™ Autophagy reagent to your cells, incubate overnight, and image—or store frozen, assay-ready cells for later use
Robust: the Premo™ Autophagy reagents are non-replicating in mammalian cells, lack observable cytopathic effect, and are suitable for biosafety level (BSL) 1

The LC3B protein plays a critical role in autophagy, and the localization of this protein to autophagosomes can be used as a general marker for formation of autophagosomes during stimulation of autophagy and/or the accumulation of autophagosomes during inhibition of autophagic flux.

Currently available non-tandem autophagy chimeras, such as Premo™ Autophagy LC3B-GFP (P36235) and LC3B-RFP (P36236), are useful for monitoring autophagosome formation, especially when combined with other chimeras or fluorescent reagents. However, the Premo™ Autophagy Tandem Sensor allows an enhanced dissection of the maturation of the autophagosome to the autolysosome. By combining an acid-sensitive GFP with an acid-insensitive RFP, the change from autophagosome (neutral pH) to autolysosome (with an acidic pH) can be visualized by imaging the specific loss of the GFP fluorescence, leaving only red fluorescence. In addition, combining the Premo™ Autophagy Tandem Sensor with the far-red emitting LysoTracker® Deep Red (L12492) allows for a three-color analysis of the autophagosomal/autolysosomal/lysosomal dynamics.

To ensure reliable, high levels of expression in a wide variety of mammalian cells, the Premo™ Autophagy Sensors utilize BacMam 2.0 expression technology. BacMam reagents (insect Baculovirus with a Mammalian promoter) do not replicate in mammalian cells, are non-cytoxic (biosafety level 1), and are ready to use. Unlike expression vectors, BacMam reagents enable titratable and reproducible expression and offer high co-transduction efficiency; therefore, multiple BacMam reagents can be used in the same cell.

Each Premo™ Autophagy Tandem Sensor RFP-GFP-LC3B Kit includes a vial of chloroquine diphosphate. Chloroquine has been demonstrated to inhibit autophagy by elevating lysosomal pH and therefore inhibiting the fusion of autophagosomes with lysosomes and preventing the subsequent lysosomal protein degradation.

LIVE/DEAD™ BacLight™ Bacterial Viability Kit, for microscopy (Invitrogen™)

The LIVE/DEAD BacLight Bacterial Viability Kit *for microscopy* is a convenient and easy-to-use kit for monitoring the viability of bacterial populations as a function of the membrane integrity of the cell. Cells with a compromised membrane that are considered to be dead or dying will stain red, whereas cells with an intact membrane will stain green.

eBioscience™ Essential Human Th1/Th17 Phenotyping Panel with Staining & Stimulation Reagents (Invitrogen™)

The Invitrogen eBioscience Essential Human Th1/Th17 Phenotyping Panel with Staining & Stimulation Reagents is designed for flow cytometry-based identity testing of Type 1 and Type 17 T-helper cells (Th1/Th17). It leverages high quality eBioscience antibodies, cell stimulation reagents (induces cytokine production), and fixation/permeabilization reagents (for intracellular markers IfN-g and IL-17) and includes a ready-to-use protocol for experimental setup, gating strategy, and analysis.

Benefits include:
Proven performance―antibodies and supporting reagents designed for flow cytometry
Highly referenced antibodies―includes one of the most referenced clones for IL-17
Turnkey kit and protocol―contains reagents and standardized gating recommendations needed to perform flow cytometry characterization

Click-iT™ Plus EdU Alexa Fluor™ 594 Flow Cytometry Assay Kit (Invitrogen™)

The Click-iT Plus EdU Alexa Fluor 594 Flow Cytometry Assay Kit provides a simplified, more robust assay for analyzing DNA replication in proliferating cells compared to traditional BrdU methods. Newly synthesized DNA is analyzed using the 532- or 561-nm laser of the flow cytometer. The Click-iT Plus formulation is compatible with standard fluorophores, including R-PE and R-PE tandems, as well as fluorescent proteins.

• Multiplexable—compatible with R-PE (and tandems) and fluorescent proteins
• Accurate—superior results compared to BrdU assays
• Fast—results in as little as 60 minutes

View selection guide for all Click-iT EdU and Click-iT Plus EdU assays for flow cytometry.

Multiplexable
Click-iT Plus Alexa Fluor 594 EdU assays can be used in conjunction with R-PE and R-PE tandems, as well as fluorescent proteins such as GFP and mCherry. The Alexa Fluor 594 labeling reagent is excited readily at either 532 or 561 nm and emits at 615 nm.

An Advanced Method Giving You Results Superior to BrdU
The most accurate method of proliferation analysis is direct measurement of DNA synthesis. Originally, this was performed through incorporation of radioactive nucleosides. This method was replaced by antibody-based detection of the nucleoside analog bromodeoxyuridine (BrdU). The Click-iT Plus EdU Flow Cytometry assay is a novel alternative to the BrdU assay. EdU (5-ethynyl-2´-deoxyuridine) is a thymidine analog that is incorporated into DNA during active DNA synthesis. Detection is based on click chemistry, which is a copper-catalyzed covalent reaction between an azide and an alkyne. In this application, the alkyne is found in the ethynyl moiety of EdU, while the azide is coupled to the Alexa Fluor dye. Standard flow cytometry methods are used for determining the percentage of S-phase cells in the population.

Mild Conditions Allow Use with Cell Cycle Dyes and Antibodies
The small size of the dye azide allows for efficient detection of the incorporated EdU using mild conditions, while standard aldehyde-based fixation and detergent permeabilization are sufficient for the Click-iT Plus detection reagent to gain access to the DNA. This is in contrast to BrdU assays that require DNA denaturation (using HCl, heat, or digestion with DNase) to expose the BrdU so that it can be detected with an anti-BrdU antibody. Sample processing for the BrdU assay can result in signal alteration of the cell cycle distribution, as well as destruction of antigen recognition sites when using the HCl method. In contrast, the easy-to-use Click-iT Plus EdU assay is compatible with cell cycle dyes. The Click-iT Plus EdU assay can also be multiplexed with antibodies against surface and intracellular markers, as well as conjugates labeled with standard fluorophores including R-PE, R-PE tandems, and fluorescent proteins (GFP and mCherry).

Quick and Simple Protocol
The Click-iT Plus EdU protocol is based on the aldehyde fixation and detergent permeabilization steps for immunohistochemical antibody labeling. However, EdU is compatible with other fixation/permeabilization agents including saponin and methanol. In just five steps you’ll be ready to analyze your cell proliferation data:

1. Treat cells with EdU.
2. Fix and permeabilize cells.
3. Detect S-phase cells with Click-iT Plus detection cocktail for 30 min.
4. Wash once.
5. Analyze.

Results can be seen in as little as 60 minutes in some circumstances, but we recommend 90 minutes for all applications.

Click-iT™ EdU Proliferation Assay for Microplates (Invitrogen™)

The Click-iT EdU Proliferation Assay for Microplates provides a more simplified and robust method for the measurement of mammalian cell proliferation compared to traditional BrdU-based methods. After incorporation of EdU (a modified thymidine analogue) into newly synthesize DNA, horse radish peroxidase (HRP) is covalently attached via the highly specific click reaction. Amplex UltraRed Reagent, an HRP substrate, is then added and the resulting fluorescence signal detected. Because of the highly specific attachment of HRP to EdU, this assay results in highly sensitive and reliable measurement of mammalian cell proliferation in a microplate format.

Click-iT EdU Proliferation Assay for Microplates features include:
Improved performance—direct conjugation of HRP to incorporated EdU improves assay sensitivity and accuracy
Increased reliability—assay format results in low well-to-well variability
Simplified protocol—results in reduced time to data; easier to follow protocol
Multiplex enabled—assay can be easily multiplexed with other cell health reagents resulting in more data per sample

Measuring cell proliferation is a fundamental method of assessing cell health, determining genotoxicity, and evaluating anti-cancer drugs. The most accurate method of measuring proliferation is by direct detection of DNA synthesis. Originally this was accomplished through incorporation of a radioactive nucleoside (e.g., 3H-thymidine). This method was replaced by antibody-based detection of the nucleoside analog bromo-deoxyuridine (BrdU). After incorporation of BrdU into the newly synthesized DNA, denaturation is required to expose the BrdU molecules to the anti-BrdU antibody. The denaturation involves harsh methods (HCl, heat, or enzymes) and is time consuming and difficult to perform consistently.

The Click-iT EdU Proliferation Assay is an alternative to the BrdU assay. The nucleoside analog EdU (5-ethynyl-2´-deoxyuridine) is added to live cells and incorporated into DNA during active DNA synthesis. The incorporated EdU contains an alkyne group which is covalently joined to the azide group present in HRP using a highly specific copper-catalyzed covalent reaction ("click reaction"). Amplex UltraRed Reagent is then added and its conversion by HRP into a highly fluorescent product is recorded using a fluorescence microplate reader (excitation: 568, emission: 585 nm). With a high extinction coefficient, good quantum efficiency, and resistance to auto-oxidation, Amplex UltraRed Reagent delivers higher sensitivity and a broader assay range than other fluorogenic or colorimetric HRP substrates. It also offers versatility through detection by either fluorescence or absorbance measurement.

Compared to anti-BrdU-based microplate proliferation assays, the Click-iT EdU Proliferation Assay uses small reaction moieties and does not require antibodies or DNA denaturation, both of which can reduce assay performance. Additionally, streptavidin-biotin binding is not required, eliminating the necessary biotin blocking steps required by cells possessing high levels of endogenous biotin.

CyQUANT™ Direct Cell Proliferation Assay (Invitrogen™)

Experience a more convenient workflow for your high-throughput cytotoxicity and proliferation screening projects with the CyQUANT® Direct Cell Proliferation Assay–no washes, cell lysis, temperature equilibrations, or radioactivity required. DNA-based assays have been shown to be among the most sensitive cell health indicators and data concordance with metabolically based assays is excellent. The CyQUANT® Direct assay is a highly sensitive and robust method to assess cell growth, viability, or compound toxicity in a range of applications, from high throughput screening to bioproduction.

Features of the CyQUANT® Direct Cell Proliferation Assay include:

• More convenient workflow for high-throughput screening–no washes, cell lysis, temperature equilibrations, or radioactivity required
• Rapid protocol, high sensitivity and dynamic range
• Measures independent of metabolic state of cells on standard fluorescent plate readers
• Multiplex assays using a spectrally distinct fluorescent or a luminescent readout

Rapid and convenient, with a large dynamic range
The CyQUANT® Direct assay is a fluorescence-based proliferation and cytotoxicity assay for microplate readers with a linear detection range from less than 100 to 20,000 cells per well in most cell types. The no-wash, homogenous format and fast add-mix-read protocol makes the CyQUANT® Direct assay ideal for HTS applications. The assay can be completed in one hour, with no washes, no cell lysis, or temperature equilibrations required. The signal is stable for several hours, affording additional work flow convenience.

How it works
The CyQUANT® Direct assay is based on a cell-permeant DNA-binding dye in combination with a masking dye reagent. Cellular DNA is highly regulated and estimates of cell numbers based on DNA are very accurate. The masking dye blocks staining of dead cells or cells with compromised cell membranes resulting in only healthy cells being stained. The CyQUANT® Direct assay, therefore, measures proliferation as well as membrane integrity, another measure of cell health. Users of metabolically based cell health assays can readily compare historical data with results from the CyQUANT® Direct assay, as concordance assays based on metabolism or energy (ATP) has been shown to be excellent. Furthermore, it is possible to multiplex with spectrally distinct fluorescent probes or luminescence assay kits because cells are not lysed. Furthermore, the fluorescent signal allows the user to switch between HTS and HCS plate readers if multiplex assays require different platforms.

Escherichia coli (K-12 strain) BioParticles™, Alexa Fluor™ 594 conjugate (Invitrogen™)

The Molecular Probes® BioParticles® product line consists of a series of fluorescently labeled, heat- or chemically killed bacteria and yeast in a variety of sizes, shapes, and natural antigenicities. These fluorescent BioParticles® products have been employed to study phagocytosis by fluorescence microscopy, quantitative spectrofluorometry, and flow cytometry.

We offer E. coli (K-12 strain), S. aureus (Wood strain, without protein A) and zymosan (S. cerevisiae) BioParticles® conjugates covalently labeled with a variety of different fluorophores (special care has been taken to remove free dye after conjugation). Unlike the fluorescence of fluorescein-labeled BioParticles® conjugates, which is partially quenched in acidic environments, the fluorescence of the Alexa Fluor®, BODIPY® FL, tetramethylrhodamine and Texas Red® dye conjugates is uniformly intense over the pH range from 3 to 10.

BioParticles Specifications:
• Label (Ex/Em): Alexa Fluor® 594 (~590/617 nm)
• Particle: E. coli (K-12 strain)
Opsonizing reagent available


Using BioParticles Products
BioParticles® conjugates are provided as lyophilized powders. There are approximately 3 x 108 E. coli or S. aureus particles per mg solid and approximately 2 x 107 zymosan particles per mg solid. BioParticles® conjugates can be reconstituted in the buffer of your choice for use in phagocytosis assays. The fluorescence of BioParticles® conjugates that are bound to the surface of the cell (but not internalized) can be quenched by ethidium bromide, trypan blue, or other quenchers. In addition to cellular applications, fluorescent BioParticles® conjugates may be effective as flow cytometry calibration references when sorting bacteria and yeast mutants. These small particles may also be useful references for light scattering studies because their sizes and shapes differ in characteristic ways.

Find More BioParticles® Products
We offer a large range of dye-labeled and unlabeled E. coli (K-12 strain), S. aureus (Wood strain, without protein A), and zymosan (S. cerevisiae) BioParticles® products. Find out about these products and their applications by reviewing Probes for Following Receptor Binding and Phagocytosis—Section 16.1 in the Molecular Probes® Handbook.

For pH-sensitive endocytosis assays, see our pHrodo™ BioParticles® conjugates.

For Research Use Only. Not for human or animal therapeutic or diagnostic use.

Vybrant™ FAM Caspase-3 and -7 Assay Kit, for flow cytometry (Invitrogen™)

The Vybrant™ FAM Caspase-3 and -7 Assay Kit employs a novel approach to detect active caspases that is based on a fluorescent inhibitor of caspases (FLICA™) methodology, essentially an affinity label. The reagent associates a fluoromethyl ketone (FMK) moiety, which can react covalently with a cysteine, with a caspase-specific amine acid sequence. For caspase-3 and -7, this recognition sequence is aspartic acid-glutamic acid-valine-aspartic acid (DEVD). A fluorescein group is attached as a reporter. The FLICA reagent is thought to interact with the enzymatic reactive center of an activated caspase via the recognition sequence, and then to attach covalently through the FMK moiety. The FLICA inhibitor is cell permeant and noncytotoxic. Unbound FLICA molecules diffuse out of the cell and are washed away; the remaining green-fluorescent signal is a direct measure of the amount of active caspase that was present at the time the inhibitor was added. This kit includes the FLICA reagent specific for caspase-3 and -7, Hoechst 33342 stain, and propidium iodide stain, which allows the simultaneous evaluation of caspase activation, membrane permeability, and cell cycle by flow cytometry.

View a selection guide for all apoptosis assays for flow cytometry.