Drug discovery is an extremely lengthy and expensive process. On average, drugs today cost more than $5 billion to develop, and take more than a decade to reach the market. Most drugs today are discovered using high throughput screening, in which hundreds of thousands to millions of trial compounds are assayed against cells to determine if they interact with a disease target. This is typically performed using at the well-level, using techniques such as fluorescence, chemiluminescence, and optical absorbance, to measure a cell population's average response to a drug. While these techniques are high throughput, they lack the ability to interrogate wells at the single-cell level which provides a much more comprehensive picture of the phenotypic drug-cell interaction than population-averaged measurement do. Flow cytometry is a well-established and widely used technique used in most areas of cell biology that provides multi-parameter, cell-level phenotypic information by measuring optical scatter and fluorescence information from individual cells in flow at a high throughput (~10,000 cells/second). While this cellular throughput is high, the sample throughput of flow cytometers is low, compared to fluorescence plate readers, for example. This speed limitation is primarily due to the serial sample handling approach employed by flow cytometers. If this speed limitation could be eliminated, flow cytometry could be used in primary screening assays. The high content phenotypic information obtained by using flow cytometry early in the drug discovery process would enable researchers to more readily understand a candidate compound's complex intra- and inter-cellular effects. This transformation in the compound screening process would increase the efficiency of drug discovery by elucidating complex drug-cell interactions at an earlier stage, ultimately reducing the overall cost and time-to-market of new drugs. Here, we propose to develop a parallel flow cytometer system using a novel optical technology, known as Fluorescence Imaging using Radiofrequency-tagged Emission, or FIRE. FIRE is a high-speed optical technique that enables a single photomultiplier tube detector to measure fluorescence or optical scatter signals from multiple points on a sample using radiofrequency-domain multiplexing. We will use a modified FIRE optical system to probe fluorescence and scatter from cells flowing in 8 parallel flow channels. This instrument will simultaneously collect flow cytometry data from 8 samples, aspirated from well plates using a multi-probe autosampler. We will design, construct, and characterize the system using standard protocols, and perform a relevant screening assay to test its performance. This system will enable a 384-well plate to be sampled in less than 2 minutes, ultimately enabling high content compound screening of more than 250,000 wells per day using flow cytometry.
An instrument capable of analyzing cells flowing in parallel is proposed for improving the drug discovery process. This instrument will enable candidate drug compounds to be screened at a rate approximately 10 times faster than the current state of the art, which will ultimately improve the efficiency at which researchers discover new pharmaceuticals.
