Methods in Molecular Biology Cover Image

We are excited to highlight the Methods Molecular Biology textbook that references the exceptional utility of FLICA for apoptosis detection ex vivo and in vivo in various tissues. FLICA allows for the detection of caspase activity. Caspases play important roles in apoptosis and inflammation.

Methods in Molecular Biology Cover Image

ICT’s FLICA assay kits are used by researchers seeking to quantitate apoptosis via caspase activity in cultured cells and tissues. For technical information from this book, read on!

Apoptosis, or programmed cell death, is a process characterized by caspase activation. Numerous cellular analysis techniques for detecting key apoptotic markers have been established to quantify apoptotic frequency within various cell populations. The approach illustrated in this chapter uses fluorescent labeled inhibitors of caspases (FLICA) and is relevant to fluorescence microscopy- including confocal imaging, as well as flow- and imaging-cytometry. Carboxyfluorescein- or sulforhodamine-labeled FLICA reagents, when exposed to live cells in vitro or in vivo, traverse the lipid bilayer and specifically label apoptotic cells.

The FLICA labeling technique is fast, easy, reliable, and can be used in combination with other cell death indicators for multiparametric analysis. Examples demonstrating the use of FLICA in conjunction with a live/dead stain (propidium iodide), detection of mitochondrial membrane depolarization, and Annexin V fluorophore conjugate-based detection of phosphatidylserine exposure on the outer cell membrane surface are presented.

FLICA stained cells can be fixed and assayed for correlation of FLICA binding with other relevant techniques requiring cell permeabilization for detection, including DNA ploidy assessment, cell cycle phase determination, and screening for DNA fragmentation and other apoptotic markers.

In comparison to Annexin V binding, the timeframe for apoptosis detection with FLICA is broader, making it a better option for detecting early stage apoptosis, before phosphatidylserine translocation occurs, and hence more accurate for quantification of apoptotic cells. FLICA’s broad utility for apoptosis detection ex vivo and in vivo in various tissues surpasses many other apoptosis biomarkers.

Reference: Darzynkiewicz Z, Zhao H, Dorota Halicka H, Pozarowski P, Lee B. Fluorochrome-Labeled Inhibitors of Caspases: Expedient In Vitro and In Vivo Markers of Apoptotic Cells for Rapid Cytometric Analysis. Methods Mol Biol. 2017;1644:61-73. doi: 10.1007/978-1-4939-7187-9_5. PMID: 28710753.

Cellular assays