Cell Health Assays for Muse® Cell Analyzer

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Confidence in your experimental results starts with knowing the upstream health status of your cells. Simple, robust viability, autophagy, apoptosis and stress assays are fundamental tools for elucidating disease mechanisms and therapeutic discovery, and establishing uniform standards of cellular performance across long-term research studies. Knowing the performance profile of your cells prior to running your bioassay can mean the difference between valid assay results and wasted reagents, lost time and discarded data. For any researchers working with cultured or primary cells, confidence in experimental results begins with knowing cell health status.

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Cell Health Assays

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Muse® Count & Viability Assay Kit

(Cat. No. MCH100102)

The Muse® Count and Viability Kit provides rapid and reliable determinations of viability and total cell count using the Muse® Cell Analyzer. Precise, accurate assessments can be made with a wide variety of cell types.

This simple no-wash, mix-and-read kit accurately determines absolute total cell counts and viability, based on differential permeabilities of two DNA-binding dyes. The nuclear dye stains only nucleated cells, while the viability dye brightly stains dying and dead cells. This proprietary combination of dyes enables the kit to distinguish viable and dead cells. Additionally, debris is excluded from results based on negative staining with the nuclear dye.

• Kit Components

  
  

  • Muse® Count & Viability Reagent (Part No. 4000-0335, 100 tests/bottle; Part No. 4000-0340, 600 tests/bottle).
  • The Muse® Count & Viability Reagent should be stored at 2-8°C and protected from light.
• Materials Recommended

  
  

  • Muse® Cell Analyzer.
  • Cell suspension.
  • Dilution buffer: Phosphate buffered saline (PBS), or equivalent balanced salt solution, pH 7.2 to 7.4. or complete growth medium.
  • Micropipettors.
  • Disposable micropipettor tips.
  • Microcentrifuge tubes with screw caps, 1.5 mL (VWR, Cat. No. 16466-030, or equivalent).
  • Muse® Count & Viability CDR (Part No. 4700-0050), optional Vortex mixer.
• Expected Results

  
  The figure on the right shows an example of results obtained using the Muse® Count & Viability Kit. Results obtained with Muse® Count & Viability Assay module using healthy Jurkat cells mixed with heat-killed Jurkat cells stained with Muse® Count & Viability Kit and acquired on the Muse® Cell Analyzer. Figure 1A shows results obtained without display of dot plots, while results in 1B show results obtained with optional dot plots shown. The statistics show the Viable Cells/mL, the % Viability, and the Total Cells/mL for the Jurkat cell sample shown. The first plot in 1B shows the Viability vs Cell Size and the second plot in 1B shows the Viability vs Nucleated cell plot.
  
  

  

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Muse® Cell Cycle Assay Kit

(Cat. No. MCH100106)

The Muse® Cell Cycle Kit can be used for measuring G0/G1, S, and G2/M phase distributions. The assay is based on the established method of whole-cell staining with propidium iodide (PI) and the single reagent contains everything you need for your assay already in solution.

• Cells in the G0/G1 phase have 2N DNA and will have the dimmest staining with PI.
• Cells in S phase are in the process of synthesizing the DNA and can have anywhere from 2N to 4N DNA and will appear as a smear between the two major peaks.
• Cells in G2/M phase will have 4N the DNA compared to the cells in the G0/G1 phase and will stain twice as brightly as the G0/G1 peak.

Using this easy and convenient kit on the Muse® Cell Analyzer will give you quick answers to your questions on cell cycle for your most important questions in cancer research, cell signaling, cell differentiation, and more.

• Kit Components

  
  

  • Muse® Cell Cycle Reagent (Part No. 4700-1495, 100 tests/bottle).
  • Muse® Cell Cycle Kit should be stored at 2-8°C and protected from light.
• Materials Recommended

  
  

  • Muse® Cell Analyzer.
  • Cell suspension.
  • Ethanol 70%.
  • Complete growth medium appropriate for your cells.
  • 12x75mm tubes.
  • Micropipettors.
  • Disposable micropipettor tips.
  • Microcentrifuge tubes with screw caps, 1.5 mL (VWR, Cat. No. 16466-030, or equivalent).
  • Muse® Count & Viability CDR (Part No. 4700-0050), optional Vortex mixer.
• Expected Results

  
  Figures 1A and 1B show typical results obtained with the Muse® Cell Cycle Reagent. Log-phase Jurkat T cells were ethanol fixed overnight and stained according to the protocol described above. Results obtained with Muse® Cell Cycle module using Jurkat cells stained with Muse® Cell Cycle Kit and acquired on the Muse® Cell Analyzer. Figure 1A shows results obtained without display of dot plots, while results in 1B show the same results obtained with optional dot plots shown. The statistics show the percentage of cells in each population, the mean intensity for each peak, and the %CV for each peak.The first plot in 1B shows the DNA content vs the Cell Size Index dot plot.The second plot in 1B shows plot shows the distribution of the cell cycle phases (G0/G1, S and G2/M) in histogram format. The DNA Histogram Results show the result for the percentage of cells in G0/G1 (M1), S (M2) and G2/M (M3).
  
  

  

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Muse® Annexin V & Dead Cell Assay Kit

(Cat. No. MCH100105)

The Muse® Annexin V & Dead Cell Kit offers a method to detect cells in various stages of apoptosis. The assay relies on the binding of fluorescently labeled Annexin V to phosphatidylserine (PS), which translocates to the outer surface of the cell membrane upon the onset of apoptosis.

The assay is a simple, mix-and-read assay that is highly sensitive and reproducible. The Muse® Annexin V & Dead Cell Kit uses a single reagent and two stains to reliably stain and differentiate live, dead and apoptotic cells. Early in the apoptotic pathway, molecules of PS are translocated to the outer surface of the cell membrane. Annexin V-PE is used to to detect PS on the external membrane of apoptotic cells. 7-AAD, because it is excluded from live and healthy cells, permeates the late-stage apoptotic and dead cells. The Muse® Cell Analyzer walks you step by step through gating up to 4 distinct cell populations: dead cells, live cells, early and late apoptotic cells.

• Kit Components

  
  

  • Muse® Annexin V & Dead Cell Reagent (Part No. 4700-1485, 100 tests/bottle).
  • Muse® Annexin V & Dead Cell Kit should be stored at 2-8°C and protected from light.
• Materials Recommended

  
  

  • Muse® Cell Analyzer.
  • Cell suspension, untreated and treated to undergo apoptosis.
  • Micropipettors.
  • Disposable micropipettor tips.
  • Microcentrifuge tubes with screw caps, 1.5 mL (VWR, Cat. No. 16466-030, or equivalent).
  • Muse® Count & Viability CDR (Part No. 4700-0050), optional Vortex mixer.
• Expected Results

  
  The figure on the right shows an example of results obtained using the Muse® Annexin V & Dead Cell Kit to stain Jurkat cells treated with staurosporine to induce apoptosis.
  
  Results obtained with Muse® Annexin V & Dead Cell module using Jurkat cells treated with 1 µM staurosporine, stained with Muse® Annexin V & Dead Cell Kit and acquired on the Muse® Cell Analyzer. Figure 1A shows results obtained without display of dot plots, while results in 1B show results obtained with optional dot plots shown. The statistics show the cells/mL in the stained cell sample and the percentages of each population. The first plot in 1B shows the Annexin V vs Cell Size and the second plot from 1B shows the Viability vs Annexin V cell plot.
  
  Events in each of the four quadrants are as follows:

  • Lower-left quadrant: viable cells, not undergoing detectable apoptosis [in V-PE (-) and Dead Cell Marker (-)].
  • Lower- right quadrant: cells in the early stages of apoptosis [in V-PE (+) and Dead Cell Marker (-)].
  • Upper-right quadrant: cells in the late stages of apoptosis or dead (by necrotic or apoptotic mechanisms) [in V-PE (+) and Dead Cell Marker (+)].
  • Upper-left quadrant: cells that have died not through the apoptotic pathway [in V-PE (-) and Dead Cell Marker (+)].

  
  
  

  

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Muse® Caspase-3/7 Assay Kit

(Cat. No. MCH100108)

The Muse® Caspase-3/7 Kit allows for the facile, rapid, and quantitative measurements of two important cell health parameters simultaneously; (1) apoptotic status based on Caspase-3.7 activation and (2) cellular plasma membrane permeabilization and cell death. The Muse® Caspase-3/7 Kit simultaneously determines the count and percentage of cells in various stages of apoptosis based on activity of executioner caspases namely, Caspase-3/7 activity in combination with a dead cell dye. The kit utilizes (1) a novel Muse® Caspase-3/7 reagent NucView for the detection of Caspase-3/7 activity and (2) a cell death dye that provides information on membrane integrity or cell death.

The assay provides relative percentage of cells that are live, in the early and late stage of apoptosis, and cell death on both adherent and suspension on the Muse® Cell Analyzer. Minimal sample preparation is required in this no-wash, mix and read assay to obtain accurate and precise results.

• Kit Components

  
  

  • Muse® Caspase-3/7 Reagent (Part No. 4700-1505, 100 tests/bottle)
  • Muse® Caspase 7-AAD (Part No. 4700-1510, 100 tests/bottle)
  • 1X Assay Buffer BA (Part No. 4700-1360, 100 tests/bottle)
  • 1X PBS (Part No. 4700-1515, 100 tests/bottle)
• Materials Recommended

  
  

  • Muse® Cell Analyzer
  • Cell suspension treated and untreated to undergo apoptosis
  • Micropipettors
  • Disposable micropipettor tips
  • Microcentrifuge tubes with screw caps, 1.5 mL (VWR, Cat. No. 16466-030, or equivalent)
  • Muse® Count & Viability Kit (Cat. No. MCH100102 (100T) or Cat. No. MCH600103 (600T)), optional
  • Muse® Cell Dispersal Kit (Cat. No. MCH100107), optional
  • Vortex mixer
  • Disposable gloves
  • 20% bleach solution
  • Guava ICF instrument cleaning fluid (Cat. No. 4200-0140), optional
  • Deionized water
  • Muse® System Check Kit (Cat. No. MCH100101)
• Expected Results

  
  Figure A and B. Jurkat cells were treated with staurosporine to induce apoptosis, then stained with the Muse® Caspase-3/7 Kit and acquired on the Muse® Cell Analyzer. Figure A shows summary data, while Figure B shows results displayed with optional dot plots. The statistics show the percentages and the concentration (cells/mL) for the gated events in each quadrant, as well as the percentage and concentration of total apoptotic cells. The first plot in Figure B shows Caspase-3/7 vs Cell Size Index and the second plot shows Caspase-3/7 vs Viability.
  
  
  

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Muse® Mitopotential Assay Kit

(Cat. No. MCH100110)

The Muse® MitoPotential Assay allows for the simultaneous measurement of 2 important cell health parameters; change in mitochondrial potential, considered an early hallmark of apoptosis, and cellular plasma membrane permeabilization or cell death. The Muse® MitoPotential Assay utilizes the MitoPotential Reagent to detect changes the mitochondrial membrane potential. A dead cell marker is also used as an indicator of cell membrane structural integrity. It is excluded from live, healthy cells, as well as early apoptotic cells. Quantitative data (percentages and concentrations) is generated for 4 populations of cells: live, depolarized, depolarized/dead, and dead cells.

Minimal sample preparation is required in this no-wash, mix –and-read assay to obtain accurate and precise results.

• Kit Components

  
  

  • Muse® MitoPotential Reagent (Part No.4700-1580) One vial containing 200 uL of MitoPotential Reagent.
  • Muse® MitoPotenial 7-AAD (Part No. 4700-1585) One vial containing 500 uL of 7-AAD.
  • 1X Assay Buffer (Part No.4700-1330) One bottle containing 100 mL of Assay Buffer
• Materials Recommended

  
  

  • Muse® Cell Analyzer
  • Cell line of interest
  • Tissue culture instruments and supplies (including 37°C incubator, growth media, detachment buffer, etc.)
  • Micropipettors
  • Disposable micropipettor tips
  • Microcentrifuge tubes with screw caps, 1.5 mL (VWR, Cat. No. 16466-030, or equivalent)
  • Vortex mixer
  • Disposable gloves
  • 20% bleach solution
  • Deionized water
  • Centrifuge
  • Guava ICF Instrument Cleaning Fluid (Cat. No. 4200-0140), optional
  • Muse® System Check Kit (Cat. No. MCH100101)

• Download Protocols and user guide

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Muse® MultiCaspase Assay Kit

(Cat. No. MCH100109)

The Muse® MultiCaspase Kit allows for the facile, rapid, and quantitative measurements of two important cell health parameters simultaneously—caspase activation and cellular plasma membrane permeabilization and cell death. Caspases (cysteinyl-directed aspartate-specific proteases) are cysteine proteases that play a central role in propagating the process of programmed cell death (apoptosis) in response to proapoptotic signals. In addition several caspases have also been found to have non-apoptotic roles in nonapoptotic functions of caspases in inflammation,mediating immunity, cell fate specification, cell survival, cell cycle regulation, cell proliferation, and cell migration. Muse® MultiCaspase Assay is a Pan Caspase assay that can detect the presence of multiple caspases (caspase-1, 3, 4, 5, 6, 7, 8, and 9). The assay simultaneously determines the count and percentage of cells with caspase activity, in combination with a dead cell dye (7AAD). It provides relative percentages of cells that are either live, exhibiting caspase activity, and dead, for both adherent and suspension cultures on the Muse® Cell Analyzer.

Minimal sample preparation is required in this no-wash, mix-and-read assay to obtain accurate and precise results. When samples are prepared with this reagent and acquired on the Muse® Cell Analyzer, the software provides the following data:

• Percentage of live, caspase+, caspase+ and dead, total caspase+, and dead cells
• Cell Concentrations (cells/mL) for live, caspase+, caspase+ and cead, and dead cells

• Kit Components

  
  

  • Muse® MultiCaspase Reagent (Part No. 4700-1530, 100 tests/bottle)
  • Muse® Caspase 7-AAD (Part No. 4700-1510, 100 tests/bottle)
  • Muse® 10X Caspase Buffer (Part No. 4700-1535, 100 tests/bottle)
  • 1X PBS (Part No. 4700-1515, 100 tests/bottle)
  • Anhydrous DMSO (Part No. 4300-0160, 100 tests/bottle)
• Materials Recommended

  
  

  • Muse® Cell Analyzer
  • Cell suspension treated and untreated to undergo apoptosis
  • Micropipettors
  • Disposable micropipettor tips
  • Microcentrifuge tubes with screw caps, 1.5 mL (VWR, Cat. No. 16466-030, or equivalent)
  • Muse® Count & Viability Kit (Cat. No. MCH100102 (100T) or Cat. No. MCH600103 (600T)), optional
  • Vortex mixer
  • Disposable gloves
  • 20% bleach solution
  • Guava ICF instrument cleaning fluid (Cat. No. 4200-0140), optional
  • Deionized water
  • Muse® System Check Kit (Cat. No. MCH100101)
• Expected Results

  
  The figures below show an example of results obtained using Muse® MultiCaspase Assay. Jurkat cells were treated with staurosporine to induce apoptosis, then stained with the Muse® MultiCaspase Kit and acquired on the Muse® Cell Analyzer. Figure A shows summary data, while Figure B shows results displayed with optional dot plots. The statistics show the percentages and the concentration (cells/mL) for the gated events in each quadrant, as well as the percentage and concentration of total caspase-positive cells. The first plot in Figure B shows Caspase vs Cell Size Index and the second plot shows Caspase vs Viability.

  
  
  

  
  
  

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Muse® Nitric Oxide Kit

(Cat. No. MCH100112)

The Muse® Nitric Oxide Kit allows for the quantitative simultaneous measurements of two important cell health parameters—changes in intracellular nitric oxide activity levels and cellular plasma membrane permeabilization or cell death. The Muse® Nitric Oxide Reagent is a membrane permeable novel reagent DAX-J2 Orange that generates a highly fluorescent product upon NO oxidation inside the cell. A dead cell marker (7-AAD) is also included in the kit as an indicator of cell membrane structural integrity and cell death. 7-AAD is excluded from live cells with intact membranes, but permeates membranes that are compromised or stressed as well as dead cells. Minimal sample preparation is required in this no-wash assay to obtain accurate and precise results on both suspension and adherent cells. The Muse® software output provides the percentage and concentration of live cells, live cells with nitric oxide activity, dead cells with nitric oxide activity, dead cells, and the total nitric oxide-positive cells (live and dead).

• Download Protocols and user guide

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Muse® Oxidative Stress Kit

(Cat. No. MCH100111)

The Muse® Oxidative Stress Kit allows for the quantitative measurements of Reactive Oxygen Species (ROS), namely superoxide radicals in cells undergoing oxidative stress. The assay provides relative percentage of cells that are ROS negative and positive in both adherent and suspension on the Muse® Cell Analyzer. Minimal sample preparation is required in this no-wash, mix-and-read assay to obtain accurate and precise results. The Muse® Oxidative Stress Kit simultaneously determines the count and percentage of cells undergoing oxidative stress based on the intracellular detection of superoxide radicals. The kit utilizes the Muse® Oxidative Stress Reagent for the detection of ROS in cells. The Muse® Oxidative Stress Reagent is based on dihydroethidium (DHE), a well characterized reagent that has been extensively used to detect reactive oxidative species in cellular populations.

Two populations of cells can be distinguished in the assay:

• ROS(-): live cells
• ROS(+): cells exhibiting ROS

• Download Protocols and user guide

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Muse® Autophagy LC3-Antibody Based Kit

(Cat. No. MCH200109)

The Muse® Autophagy LC3-antibody based kit provides a quantitative solution for the study of autophagy. This kit contains two key detection reagents to help facilitate the monitoring of lipidated LC3-II:

1. The use of selective permeabilization solution discriminates between cytosolic LC3 from autophagic LC3 by extracting the soluble cytosolic proteins, while protecting LC3 which has been sequestered into the autophagosome;
2. Since autophagy is a constitutive cellular degradation process, the use of an autophagy detection reagent (Autophagy Reagent A) will prevent the lysosomal degradation of LC3, allowing its quantification by flow cytometry.

Muse® Autophagy LC3-antibody based kit includes an anti-LC3 mouse monoclonal antibody conjugated to Alexa Fluor®555, used to measure and track the levels of LC3 within the cell. The anti-LC3 Alexa Fluor®555 conjugated antibody and autophagy enabling reagents have been carefully evaluated to ensure optimal performance, alleviating the need for any additional validation of the kit reagents. This kit contains reagents and buffers to provide researchers a complete solution for autophagy analysis. Data generated using the Muse® Cell Analyzer along with the corresponding Muse® software module provides statistical values measuring:

• Mean Autophagy Value (for both control and test samples)
• Autophagy Induction Ratio (Test sample fluorescence relative to control)
• Percentage of cells with increased autophagy (Test sample versus control)

• Download Protocols and user guide

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Muse® RFP-LC3 reporter Autophagy Assay

(Cat. No. MCH200110)

The Muse® RFP-LC3 Reporter Autophagy Assay kit includes one immortalized RFP-LC3 reporter cell line to measure and track the levels of LC3 within the cell. The RFP-LC3 reporter cell line provided in the kit has been carefully optimized along with the corresponding reagents to ensure optimal performance. This kit contains reagents and buffers to provide researchers a complete solution for autophagy analysis. The autophagy detection reagents and cell line have been optimized together to ensure the ability to measure and discriminate between cytosolic and lipidated LC3 to accurately measure the autophagic process. To assist in autophagy study, U20S (human osteosarcoma) cells are stably transfected, which are suitable for quantitative and higher throughput analysis of autophagy. Data generated using the Muse® Cell Analyzer along with the corresponding Muse® software module provides statistical values measuring:

• Mean Autophagy Value (for both control and test samples)
• Autophagy Induction Ratio (Test sample fluorescence relative to control)
• Percentage of cells with increased autophagy (Test sample versus control)

• Download Protocols and user guide

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