Assays for Targeted Protein Degradation

The activity of small molecule Degraders can be readily profiled using high throughput Simple Western systems. The highly reproducible and quantitative results generated by Simple Western allow for easy determination of DC₅₀ values (the concentration of Degrader that induces 50% degradation of the target protein) within a short time frame. Fully automated Simple Western assays allow separation and analysis of proteins by size (or charge) from 2 kDa to 440 kDa in as little as 3 hours. Western blotting antibodies may be used for detection, as well as any antibody in the large and expanding database of Simple Western validated antibodies.

Bio-Techne provides a wide range of primary antibodies validated for western blot and Simple Western assays.  We also offer enzyme- and fluorochrome-conjugated secondary antibodies  as well as antibodies for the most commonly used epitope tags for the detection of tagged proteins by  Western blot. We offer a variety of size options, including bulk and sample sizes to ensure that you can find what you need for your research.

Controls should be included to assess whether target degradation and pharmacological effect are ‘on-mechanism’ i.e. mediated by the ubiquitin-proteasome system (UPS). This can be achieved via co-treatment with a proteasome inhibitor, such as Bortezomib (#7282), which should block Degrader-mediated target protein degradation. A useful additional control is co-treatment with a NEDD8-activating enzyme inhibitor, for example MLN 4924 (#6499) to confirm that an active E3 ligase is involved in degradation.

RNAscope in situ hybridization technology is a powerful tool that allows you to spatially map and quantify specific target mRNAs in intact cells and tissues. Detect up to 12 RNA targets simultaneously using the RNAscope HiPlex assay or combine the assay with IHC or IF on the same slide to investigate mRNA and protein co-expression. Catalog probes are available for over 23,000 targets in 140+ species and can also be made-to-order.  Discover how your Degrader affects downstream gene expression using RNAscope.

Proteome Profiler Antibody Arrays are a simple, high throughput and cost-effective tool for early-stage analyte profiling.  Our antibody arrays have superior specificity, low background noise and no cross-reactivity means that you can count on this assay for clear and consistent data. Use our antibody arrays as part of your PROTAC® assay cascade to explore the downstream consequences of Degrader-induced protein knockdown.

Degradation of targets can have cytotoxic effects and can interfere with transcription, resulting in DNA damage. Bio-Techne offer CometAssay, a sensitive technique for measuring DNA damage in single cells. Can be used to detect single- and double-stranded DNA breaks or to detect double-stranded DNA breaks only. For more information about CometAssay and how it can be used, please browse the products with the link below.

The viability and proliferation of a cell population after treatment with a Degrader can help to evaluate the toxicity and effectiveness of a series of candidate Degraders. Bio-Techne provides a range of assay kits and reagents to determine how your candidate Degrader affects cells.

With the Annexin V-FITC Apoptosis Detection Kit you can detect and measure apoptotic cells to quantify the extent of cell death resulting from targeted protein degradation. 

A ready-to-use solution for high throughput cell viability and proliferation assays.

Dr Gary Allenby, CSO at  Aurelia Bioscience, shares how their research team uses Simple Western systems to rapidly and reproducibly quantify protein knockdown following treatment with Degraders.

Learn from Protein Simple how fully automated Simple Westerns can be used to profile degradation of cereblon neosubstrates using cereblon-recruiting Degraders and IMiDs.