Screening tools for CBL activity from BPS Bioscience

CBL-B and c-CBL are E3-ubiquitin ligases that act as intracellular checkpoints by targeting proteins for degradation via the proteasomal pathway. CBL-B specifically functions as negative effector of T cell activation by inducing down regulation of the T cell receptor. A mutation in the CBL-B gene has been linked to type 1 diabetes and potentially other autoimmune diseases. C-CBL interacts with a large number of receptor tyrosine kinases involved in cell signaling and activation, targeting them for degradation.

BPS two options of CBL-B and c-CBL TR-FRET assay kits can measure protein auto-ubiquitination or provide additional insights on polyubiquitination using their intrachain assay kits.

TR-FRET assay kit advantages

• Homogeneous assays, ideal for high-throughput applications
• Available in 384-well plate format
• All components are included with the kits
• Use with any plate reader capable of measuring TR-FRET

TR-FRET assay kits for CBL-B and c-CBL

These TR-FRET assay kits are designed to measure c-CBL or CBL-B auto-ubiquitination activity in a homogeneous 384 reaction format. They utilize biotin-labeled ubiquitin and a terbium-labeled anti-GST antibody that targets GST-tagged c-CBL or CBL-B to complete the TR-FRET pairing. These FRET-based assays require no time-consuming washing steps, making it especially suitable for high throughput screening applications to determine effective inhibitors of c-CBL and CBL-B. The limitation of these kits is that they do not differentiate between mono-ubiquitination or polyubiquitination.

Tyro3 ubiquitination intrachain TR-FRET assay kits

The c-CBL and CBL-B-driven Tyro3 ubiquitination intrachain TR-FRET assay kits are sensitive, high-throughput screening (HTS) TR-FRET assay kits, designed to measure c-CBL or CBL-B E3 ligase activity in a homogeneous 384 reaction format. They utilize a Europium cryptate-labeled Ub (donor) and a Cy5-labeled Ub (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 high throughput applications as well as real-time analyses of polyubiquitination. Of note, the assay kit does not detect mono-ubiquitination.

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