In vitro biochemical assay development can be a long and expensive process, consuming months and thousands of dollars.1 Finding an off-the-shelf solution that’s been proven in drug discovery can help remove some of the painstaking labor required for enzymatic assay development.
While there is a wide range of biochemical assays for high throughput screening commercially available, each assay method is unique. Some have been developed for individual targets, while others can be used with a broader assortment.
Universal activity assays can help simplify the process as researchers work on multiple targets within an enzyme family. These assays work by detecting a product of an enzymatic reaction common between various targets. One example is studying a variety of kinase targets with the same universal ADP assay.
Universal activity assays detect the products produced by the enzyme allowing multiple targets to be studied with the same assay. Above is the fundamental assay concept for the Transcreener platform. Using competitive direct detection with various antibody and tracer modifications provides multiple fluorescent formats such as FI, FP, and TR-FRET.
A Universal Biochemical Assay Provides Options
Sometimes universal assays are the only commercially available option for challenging targets. And since they measure the products of an enzymatic reaction, it’s easy to determine enzymatic activity. This information allows the scientist to learn how a compound modulates the target protein’s enzymatic properties. Popular data points include IC50 or EC50, both of which can be determined quickly to accelerate SAR.
Some assays like Transcreener were designed with a simple mix-and-read format. Once the concentration of detection reagents is established just run the enzyme reaction, add the detection reagents, and read the plate. This configuration is amenable to high throughput screening. Fewer steps help simplify automation and produce robust results.
Mix-and-read assays are great for HTS and lead discovery due to their simple yet robust nature. After the enzyme reaction is complete, add the detection reagents, incubate, and read the plate.
Achieving Assay Success
Once an assay has been used, it becomes inherently familiar to the scientist. This familiarity can make it easier to develop assays for other targets at a much more rapid pace. There will always be small modifications based on the target, buffer conditions, and substrate. If the assay system is robust, adapting new targets to the system can be done with limited development.
Assay validation can be challenging, knowing the expected characteristics for robust data such as Z’ can simplify. Although there is always some method validation required prior to any screen, HTS ready biochemical assays for enzyme targets can help mitigate poor assay performance. This moves thoughts from “Is my assay working?” to “What is the data telling me?”
Instrumentation is also critical to assay success. Once a universal platform has been introduced, the correct instrument settings have been determined. Understanding a plate reader, using the right assay plate, and familiarity how an assay’s readout is achieved are all critical factors to the project. Once settings are optimized for speed and accuracy, the same configuration can be used for new targets and thus simplifying future development.
Simplify Biochemical Assay Development and Follow Up Studies
As new assay techniques are introduced commercially, technology affords the opportunity to accelerate drug discovery and save time in the early stages of hit-to-lead. Biochemical assays can then be used in follow up studies, including MOA, and SAR, making the assay a multi-tool of drug discovery.
An assay with a proven track record can eliminate much of the development process by providing an out-of-the-box solution. So when looking to accelerate your biochemical assay development, consider proven universal assay kits. Many can be used for your protein target of interest, and provide a robust but straightforward option.
- Assay Guidance Manual [Internet]. Sittampalam GS, Coussens NP, Brimacombe K, et al., editors. Bethesda (MD): Eli Lilly & Company and the National Center for Advancing Translational Sciences; 2004-2019.
- Principles of early drug discovery JP Hughes, S Rees, SB Kalindjian, KL Philpott Br J Pharmacol. 2011 Mar; 162(6): 1239–1249. doi: 10.1111/j.1476-5381.2010.01127.x