MTT and MTS assays indirectly measure cell viability through measurements of mitochondrial metabolic rate. Trypan blue exclusion assays, while simple and inexpensive, require a large number of cells, are time consuming, and are often influenced by user bias 1. Traditional cell-based assays that are used to measure cell viability include: trypan blue exclusion assays, MTT/MTS, and Annexin V-FITC flow cytometry staining. Here, we present a novel method for utilizing quantitative imaging to simultaneously analyze heterocellular phenotypes in response to relevant, co-occurring environmental stimuli. However, the limitations of these standard assays have become increasingly apparent with the discordance between in vitro and clinical data and the failure of most drugs to receive FDA approval. This protocol allows one to identify subpopulations within a co-culture system and characterize the particular response of each to external stimuli.Ĭell-based assays have been a workhorse in basic research and drug development settings. We demonstrate this application using response to drug in a cancer model however, it can easily be applied more broadly to other physiological processes. However, in some cases, cell populations may be difficult to identify and quantitate based on complex cellular features and will require additional troubleshooting we highlight some of these circumstances in the protocol. This platform allows for a more comprehensive characterization of subpopulation response to perturbation while utilizing shorter time, smaller amounts of reagents, and lower likelihood of error than traditional cell biology assays. We highlight our ability to distinguish between cell types based upon either fluorescence intensity or inherent morphology features depending on the application. morphology changes, proliferation, apoptosis) of heterogeneous cell populations to changes in environmental stimuli. Using a quantitative imaging-based approach, we present a high-content protocol for characterizing the dynamic phenotypic responses ( i.e. Existing cell biology techniques often do not allow for accurate interpretation of this interplay. Cellular processes are complex and result from the interplay between multiple cell types and their environment.
0 kommentar(er)
