Optimized, automated assays for HCS

The later stages of apoptosis are characterized by changes in nuclear morphology, including chromatin condensation, degradation of nuclear envelope, and DNA strand breaks. TUNEL assays are the most widely used method to detect apoptotic DNA fragmentation in tissue samples and also work well for cultured cells. This assay uses a nuclear stain to identify cells and a TUNEL label to measure DNA strand breaks. The Invitrogen Click-iT TUNEL Alexa Fluor Imaging Assay kits are fast and efficient and offer precise, quantitative data even with high levels of apoptotic cells. The kits are optimized for HCS and provide a choice of three wavelength options to aid in multiplexing with other cellular measurements.

Typical apoptosis-TUNEL assay

Fixed-cell assay protocol.

Imaging mode

  • Widefield or confocal
  • 20x magnification

Automatically measured properties

  • Fluorescence intensity, morphology, and count values for each object
  • Fluorescence intensity, morphology, and count values of spots in different cell regions
  • Average fluorescence intensity difference and ratios between different regions for each cell
  • Intensity and count ratio between channels within different regions for each cell

TUNEL assay images

Two-panel HCS image of nonapoptotic and apoptotic cells

HeLa cells treated to induce apoptosis and imaged. Untreated (Left) and staurosporine-treated (Right) HeLa cells stained using the Invitrogen Molecular Probes Click-iT TUNEL Alexa Fluor 594 Imaging Assay, for Microscopy & HCS and Invitrogen Molecular Probes Hoechst 33342. Cells were imaged using a Thermo Scientific HCS instrument, and fluorescence intensities and count ratios were measured using Thermo Scientific HCS Studio Software.

Sample experimental data

Four-panel graph of data for different angiogenesis assays

Dose-response plots for a variety of compounds and cell types using Invitrogen Molecular Probes Click-iT TUNEL Alexa Fluor 647 to measure apoptosis. Responses were measured automatically using Thermo Scientific HCS platform with Thermo Scientific HCS Studio 2.0 Cell Analysis Software and plotted using GraphPad Prism software.