Visualizing Apoptosis in 3D Cell Cultures

Visualizing Apoptosis in 3D Cell Cultures

Apoptosis, or programmed cell death, is an important cellular mechanism that is critical in development and tissue homeostasis.  Apoptosis also plays a role in cancer biology.  For example, visualizing apoptosis in response to treatments is one way to characterize potential therapies.

While 3D cell cultures are ideal for mimicking a tumor microenvironment, obtaining apoptosis data after treating the cultures can prove difficult.

To overcome this difficulty and to make apoptosis data accessible to researchers using Cell-Mate3D™ matrix, we optimized the commonly used Invitrogen™ CellEvent™ Caspase-3/7 Green Detection Reagent that enables quick and reliable imaging of apoptotic cells in culture.

Cell-Mate3D™ cultures were setup, treated, and analyzed as followed:

  • AU565 human breast cancer cells (HER2-positive) were embedded into the Cell-Mate3D™ matrix.
  • One sample was left untreated and an equivalent sample was treated with 100 μM Taxol for 12 days.
  • Samples were stained with 15 μM CellEvent™ Caspase-3/7 reagent (three times the recommended concentration).
  • Green-fluorescent apoptotic cells were clearly seen in the Taxol-treated sample by inverted confocal microscopy.
  • Caspase staining appears brighter in Taxol treated cultures compared to non-Taxol treated cultures.

Figure 1. Detection of apoptotic cells in the Cell-Mate3D™ matrix using CellEvent™ Caspase-3/7 Green reagent. AU565 breast cancer cells were embedded in the Cell-Mate3D™ matrix and cultured for 12 days. Cells were untreated (LEFT) or treated (RIGHT) with 100 μM Taxol for 12 days to induce apoptosis. Cells were then incubated with the Invitrogen™ CellEvent™ Caspase-3/7 Green Detection Reagent (15 μM) for 30 min. to label apoptotic cells with green fluorescence, counterstained with the Invitrogen™ NucBlue™ Live ReadyProbe™ Reagent, and imaged using an inverted confocal microscope at 20x magnification.

Reagents Used

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