Toxicology profiling is a key part of drug discovery and safety as it provides a crucial means of ranking compounds for consideration in drug discovery. Traditionally toxicity testing has been carried out at late-stage drug discovery in animal models.
It is now recognized that early identification of on- and off-target drug effects results in more timely elimination of drugs, which serves to reduce drug development costs. Furthermore, high-throughput in vitro cell-based assays, which have the advantage of speed, reduced reagent usage, and reduced information content, are becoming a more accepted format.
Cell-based assessment of toxicity is best served by a multiparametric approach in order to establish both phenotypic and mechanistic information. There are a number of key cytotoxic phenomena that can be measured including plasma membrane integrity, DNA damage, DNA synthesis, mitotic arrest, and cytoskeletal disruption.
This tutorial describes the use of TTP LabTech’s Acumen® eX3 microplate laser scanning cytometer to investigate cytotoxicity using a number of high-throughput, cell-based, ready-to-use assay kits and reagents from Invitrogen, part of Life Technologies.
Laser-scanning cytometry, unlike automated fluorescence microscopy, has the ability to rapidly detect and simultaneously quantitate multiparametric data from individual cells. The Acumen eX3 laser scanning cytometer uses multiple scanning lasers (405 nm, 488 nm, 633 nm) for fluorescence excitation, in conjunction with highly sensitive photomultiplier tubes, for the detection of fluorescently labelled cells, hence offers true multiplexing with a theoretical maximum of 12 data-collection channels.
The laser-scanning approach enables whole-well scanning, which results in the generation of improved and more robust data sets. This instrument can perform optimal signal acquisition in plate densities up to 1,536 wells with scan times of <10 minutes/plate.