| Control of cell cycle progression and initiation of the
apoptotic cascade are tightly regulated processes
complicated by the myriad of signaling pathways
contributing to their regulation. Functional assays
capable of interrogating both processes in live cells are
critical for target validation and compound optimization.
In order to improve screening efficiency, we have
developed the microcapillary-based Guava EasyCyte
system, which, when used in conjunction with Guava’s
suite of functional assays, provides an ideal platform for
cellular analyses. For over 200 test compounds analyzed,
the following parameters were measured: (1) Caspases
3/7, and 8 enzyme activity; (2) changes in mitochondrial
membrane potential; (3) translocation of phosphatidylserine
residues to the cell surface; (4) membrane
permeability; and (5) cell cycle distribution. The simultaneous
interrogation of Caspase activity and cell cycle
progression, along with the information regarding the
relative frequencies of treated cells in the early, mid- or
late stages of apoptosis offers a powerful tool for both
“hit” compound identification and endpoint analysis.
In addition, this strategy provides a more detailed
understanding of the mechanism of action both at the
level of the individual cell and the overall cell population. |
