The CBL-B-driven Tyro3 Ubiquitination Intrachain TR-FRET Assay Kit is a sensitive high-throughput screening (HTS) TR-FRET Assay Kit, designed to measure CBL-B E3 ligase activity in a homogeneous 384 reaction format. It utilizes a Europium-labeled Ubiquitin (donor) and a Cy5-labeled Ubiquitin (acceptor) to complete the TR-FRET pairing. Since both the TR-FRET donor and acceptor are incorporated into poly-ubiquitin chains, this FRET-based assay requires no time-consuming washing steps, making it especially suitable for HTS applications as well as real-time kinetics analyses of polyubiquitination. Of note, the assay kit does not detect mono-ubiquitination.

Alternatively, under the same experimental conditions the AXL kinase (BPS Bioscience #40180) can be used instead of Tyro3 as C-CBL ubiquitination substrate.

Figure 1. CBL-B-driven Tyro3 ubiquitination intrachain TR-FRET Assay Kit schematic