An Overview of High Throughput Screening

         

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What Is High Throughput Screening?

High throughput screening, or HTS, is the automated testing of thousands to millions of biological, genetic, chemical, or pharmacological samples. Originally developed in the early 1990s, high throughput screening is now widely used in both basic and translational research, and it usually relies on liquid handling devices, robotics, plate readers, and instrument control and data processing software.¹ Researchers use high throughput screening to quickly identify candidates that they want to focus on for further studies. 

General Steps and Considerations for High Throughput Screening

Miniaturization, automation, and quick assay readouts are key technical considerations for setting up laboratory workflows for high throughput screening.² Miniaturization helps reduce assay reagent amounts, automation saves researchers’ time and guards against pipetting errors when dispensing reagents in the microliter to nanoliter range, and fast and sensitive readers ensure that researchers can test thousands of samples quickly. High throughput screening workflows integrate four general components: sample and library preparation, assay readout, robotic workstations and automation, and data acquisition and handling.

Sample and Library Preparation

Researchers typically prepare their high throughput screens by adding samples and other assay components to 96- to 3456-well microplates. The type of microplates they use depends on the nature of their samples and the assays they wish to perform, a critical consideration for quality control (QC) measures.³ Sample libraries are usually stored in stock plates and are often transferred by automated pipetting stations to the assay plates. Researchers can generate these libraries with samples created in their laboratories or by purchasing commercially available compounds.

Assay Readout

Researchers use plate readers or detectors to assess the results of chemical reactions in each well. A wide range of detector systems is commercially available to read fluorescence, luminescence, absorption, and other specific parameters for a high throughput assay.⁴ Once set up, these readers need minimal or no human input. 

Robotic Workstations and Automation

In high throughput screening, an integrated automation system typically takes care of sample and reagent transfer, addition, mixing, and the final readout. As part of an automation system, liquid handlers dispense and mix reagents for chemical reactions. Robots can also be used to transfer the microplates between different locations, such as from pipetting stations to detectors, within the workstation to further automate the process. An HTS system can prepare and analyze many plates simultaneously, with ultra-high throughput screening systems capable of analyzing over 100,000 samples in a single day. 

See Also “The Latest in Lab Automation”

Data Acquisition and Handling 

Because high throughput screening can generate an enormous amount of data, researchers use various software packages to process and analyze the data and also for QC. Still, data analysis and interpretation remain one of the most challenging aspects of the entire process. 

Hits

The samples or compounds that a high throughput screen identifies as promising candidates for further exploration are called hits, and researchers need to determine the specific identification parameters that are suitable for their assay.⁵ For example, they may choose a cutoff value from detection readouts to filter out the samples that fail to reach that threshold, or they may choose the samples with the highest one percent activity in an assay. 

QC Measures

QC measures ensure the validity of a high throughput screen’s results, and the absence of QC can lead to wasted time and resources. Broadly, QC measures fall under two categories: plate-based controls and sample-based controls.⁶ Plate-based controls characterize the performance of a plate’s controls and can identify issues in an assay, such as pipetting errors and the edge effect, which is caused by the evaporation from wells at the plate’s edge. Sample-based controls characterize variability in biological responses or sample potencies. For example, the minimum significant ratio, a widely used QC tool, measures assay reproducibility and characterizes the potencies of either controls or samples between assay runs. 

         

High Throughput Screening in Drug Discovery

High throughput screening has several applications in drug discovery processes.⁷ In a recent study published in the Journal of Biological Chemistry, researchers used high throughput screening to identify small molecules from an FDA-approved drug library that specifically bound to cardiac MyBP-C (cMyBP-C).⁸ cMyBP-C interacts with myosin and actin to modulate cardiac muscles, and these molecules may be useful for treating patients with heart failure. 

Researchers also use high throughput screening in precision medicine programs to choose appropriate drugs based on patients’ tumor characteristics. In a study published in Cancer Research, researchers screened a library of 126 anticancer drugs on samples collected from children with high-risk cancer to determine the optimal treatment strategy.⁹

High Throughput Screening in Other Research Fields

Though high throughput screening was originally developed by pharmaceutical companies to find suitable drug targets, it is now used in other areas of research. In a recent study published in Nature Communications, researchers developed a high throughput testing method for detecting hepatocellular carcinoma from liquid biopsies.¹⁰ In another study published in Disease Models & Mechanisms, researchers used Caenorhabditis elegans to detect potentially toxic compounds in a high throughput assay.¹¹ This method allowed them to study the various effects of these compounds, as well as many anti-infective molecules, on these nematodes over two lifecycles.