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Apoptosis Assays for Muse® Cell Analyzer

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Programmed cell death may proceed via the intrinsic or extrinsic pathways. Muse® kits were designed to help researchers not only detect and quantify apoptosis, but to also characterize it by examining phosphatidylserine translocation, mitochondrial membrane potential (ΔΨm), and caspase activation. Muse® apoptosis kits allow the study of early, mid, and late apoptosis, and also a pan-caspase assay for broad apoptosis activity.

Apoptosis Assays

Muse® Annexin V & Dead Cell Assay Kit

(Cat. No. MCH100105)

This apoptosis assay follows an easy protocol to detect cells in various stage of apoptosis
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.
    Merck:/Freestyle/BI-Bioscience/Cell-Analysis/Muse/assays/muse_caspase.gif


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Muse® Mitopotential Assay Kit

(Cat. No. MCH100110)

Cell Health experiments are made easier with Muse Cell Analyzer and the Mitopotential assay kit
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)

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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
The Multicaspase Assay kit determines the count and percentage of cells with caspase activity, in combination with a dead cell dye (7AAD)
  • 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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