Real-Time Cell Death Imaging and Analysis

Scientific innovation is dependent upon the power of observation and the strength of conclusions that are made from those observations.  In the field of in vitro biology, a majority of physiologically relevant outcomes adhere to dose- and exposure-dependent factors.  Unfortunately, data collection for traditional cell-based experiments often occurs at arbitrary but convenient endpoints and with inadequate or poorly informative tools.  The resulting data typically lack the appropriate kinetic resolution to fully answer details of the cause-and-effect relationship.  Because biological pathways and processes are already inherently complex, a new, more efficient screening paradigm is sought which allows scientists to formally examine the important parameters of “when” and “how” to better characterize a given response.

Real-time assay reagents and the multi-modal instruments which capture their signals are uniquely capable of directly addressing the temporal parameter associated with a biological response.  Incremental but important advancements in both areas have led to synergistic methodological improvements that now eclipse the mere sum of their parts.  In this work we showcase how real-time bioluminescence and fluorescence measures can be repeatedly collected over a treatment exposure using an environmentally controlled instrument capable of both intensity measures and multiple concurrent imaging functions.  We chose to demonstrate this enabling functionality in two cell models dosed with serial dilutions of two histone deacetylase inhibitors (HDACi) to study the dose-dependent progression of apoptosis and cell death. The data reveal that A549 and HepG2 respond differently with respect to the kinetics, potency, and magnitude of apoptotic susceptibility by the HDACi romidepsin and panobinostat.  Further, we detail how intensity and imaging measures produce complementary and more complete data sets associated with the cell death phenotype.  Adoption of this integrated workflow promises to be important for identifying and optimizing pharmacological agents useful for exploration of basic science and drug discovery.