Category: Automation and High-Throughput Technologies

1309-E - Flow Cytometry Multiplexing and its Use in Detailed Characterization of Cell Health

Wednesday, February 7, 2018
11:30 AM - 12:30 PM

High throughput screening (HTS) is an extremely effective method that allows researchers to identify putative active compounds for therapeutics. Flow cytometry can also be a useful platform for HTS with the added benefit of cell-by-cell analysis. Flow cytometry is a fast and sensitive platform that can analyze thousands of cells per sample, aiding in the study of protein-protein interactions, metabolic activities, as well as DNA content in a single or multi-parametric assay format. For this study, we screened several compounds within a library and compared the results using a plate reader and a flow cytometer equipment with an plate auto sampler. Jurkat and Ramos cells were cultured under standard conditions and at either hypoxic (1%) or hyperoxic (≥19%) oxygen levels for varied lengths of time; 24, 48, or 72 hrs. Membrane integrity and metabolic activity were measured as an output for evaluating cellular viability. Post-screening analysis was implemented to establish a drug dose-response and apparent EC50 determination for “hits” from the compound library. Screening analysis identified target compounds that were taken on to further tests using different mechanisms of action to assess compound toxicity. Results indicate that compound “hits” and potency differed in the screen depending on cell type, the mechanistic readout analyzed, and mode of readout used to perform the HTS experiments.

Flow cytometery has the added benefit of multi-parametric analysis, allowing for several cell health parameters to be surveyed in a single sample. Monitoring multiple read-outs at one time within a single sample allows for small sample volume preparation, less sample numbers, and allows for further analysis on understanding cell health. Here we show that cell cycle state, early apoptosis, mitochondrial health, and cell viability can be monitored as a multiplexed readout in one sample across different compound treatments. To screen a variety of drugs within one experiment, a plate auto-sampler was used for high throughput analysis, allowing for shorter run time, large data set acquisition, and small sample requirement. Monitoring multiple cellular parameters at one time can help researchers understand the complex nature of cellular toxicity due to a particular compound treatment.

Leticia Montoya

Scientist III
Thermo Fisher Scientific
Eugene, Oregon

Leticia Montoya is a chemical biologist, with a Doctorate in Chemistry from University of Oregon, and has experience in chemical approaches towards analyte detection in biological systems, development of immunoassay kits against metabolic and signaling targets, and fluorescence microscopy. She possesses a wide variety of advanced techniques, including analytical chemistry, organic synthesis, and molecular biology.

Leticia works at Thermo Fisher Scientific as a Product Development Scientist for the Development Biology R&D Team. She develops products and solutions for detection of cellular targets and functions using flow cytometry and other cell-based assay platforms.