Category: Assay Development and Screening
Cell death occurs throughout the life of an organism, and this is critical for developmental plasticity and organismal health, in part by eliminating unneeded and unhealthy cells in a timely and effective manner. However, dysfunctional programmed cell death leads to diseases such as cancer, neurodegeneration, and ischemic damage. Two classic pathways of cell death are apoptosis and necrosis. Apoptosis is an active, programmed process of autonomous cellular dismantling that avoids eliciting inflammation. Necrosis is a more passive process with uncontrolled release of inflammatory cellular content. Healthy cells respond to death-inducing stimuli by imitating a variety of molecular pathways leading to cell death. Completion of the proper pathway is a critical cellular function to ensure that the appropriate outcome is ultimately achieved.
The quantification of the cell death response is an integral component of exploring cell biology, responses to cellular stress and performing high-throughput drug screens. Classic methods of cell death detection include flow cytometry, which requires extensive handling of cells and only provides end-point data. Kinetic imaging, in contrast, is a critical application for studying dynamic biological processes in real time. Kinetic analysis of cell death analysis allows for sensitive, real-time determination of the accumulation of both apoptotic and necrotic events within the cellular population.
Here we present the use of apoptosis and necrosis dyes in combination with automated kinetic imaging to quantitatively assess the effects of known inducers of cell death in multiple cell lines. We use high contrast label free bright field imaging to assay for total number of cells and cellular dyes to label both apoptotic and necrotic cells concurrently. This allows for determination of percent apoptosis and necrosis in each population over 48 hours of drug treatment.
Sarah Beckman– Principal Scientist, BioTek Instruments, Winooski, VT
Sarah Beckman is a Principal Scientist at BioTek Instruments, Inc. She holds a PhD in Cellular and Molecular Pathology from the University of Pittsburgh. Sarah’s graduate school work utilized microscopy and image analysis to better understand methods to optimize stem cells for cardiac and skeletal muscle regeneration. As part of her postdoc at Cincinnati Children’s Hospital Medical center Sarah performed live imaging with zebrafish in order to study macrophage migration during development. Current projects in the lab focus on object based spot counting analysis and zebrafish imaging.