Revvity, 64BDIAPB2PEH, HTRF cIAP1B2 Binding Kit, 10,000 Assay Points

Overview
A fast and easy way to identify and characterize new binders to cIAP1 protein targeting BIR2 domain.
cIAP1, also named BIRC2, is a member of the Inhibitor of Apoptosis Proteins (IAP) family. IAP proteins are involved in multiple biological processes, such as innate immunity and play an important role in apoptosis inhibition.
cIAP1 overexpression has been associated with cancer resistance, making it an attractive target in cancer therapy, where several therapeutic strategies are investigated such as SMAC mimetics.
Moreover, cIAP1 displays E3 ligase activity and leads to targeted proteins ubiquitination and their subsequent degradation, such property being exploited through Proteolysis-targeting chimeras (PROTACs) strategy. Indeed, SNIPERs molecules (Specific and Non-genetic inhibitor of apoptosis protein [IAP]-dependent Protein Erasers) can induce the degradation of both the targeted proteins and IAPs and is expected to harness cancer cells killing.
Therefore, new compounds targeting cIAP1 which exhibit dual role i) inhibition of cIAP1 anti-apoptotic function and ii) induction of targeted protein degradation represent a promising therapeutic approach.
cIAP1, like other IAP family proteins, contains BIR domains which interact with IAP-binding motif of partners such as caspases. This interaction has been suggested to control pro- and anti-apoptotic activities.
cIAP1 compounds characterization on BIR2 and BIR3 domains enables accurate profiling and selectivity studies.
How it works
Assay principle
The HTRF cIAP1 BIR2 Binding Kit is a competitive assay format which uses Streptavidin-d2 /LCL161-Biotin as cIAP1 BIR2 ligand, a GST tagged human cIAP1 BIR2 binding domain, and an anti GST Cryptate-labeled antibody. cIAP1 binding compounds compete with the LCL161-Biotin Ligand and thereby prevent FRET from occurring.

 

Assay protocol
The cIAP1 BIR2 binding domain assay can be run in a 96- or 384-well low volume white plate (20 µL final). As described here, samples or standards are dispensed directly into the assay plate. The human GST-tagged cIAP1 BIR2 binding domain is then added, followed by the dispensing of the HTRF reagents: the anti GST antibody labeled with Terbium cryptate, and the LCL161-Biotin/ Streptavidin labeled with d2. The reagents labelled with HTRF fluorophores may be pre-mixed and added in a single dispensing step. No washing steps are needed. The protocol can be further miniaturized or upscaled by simply resizing each addition volume proportionally. Application-Protein-Protein Interaction
Automation Compatible-Yes
Brand-HTRF
Detection Modality-HTRF
Product Group-Kit
Sample Volume-5 µL
Shipping Conditions-Shipped in Dry Ice
Target Class-Binding Assay
Technology-TR-FRET
Therapeutic Area-Inflammation Neuroscience Oncology & Inflammation
Unit Size-10,000 assay points