Revvity, 64XIAPB3PEH, HTRF Human XIAP / BIR3 Binding Kit, 10,000 Assay Points

Overview
A fast and easy way to identify and characterize new binders to XIAP protein targeting the BIR3 domain.
X-linked inhibitor of apoptosis protein (XIAP) protein belongs to the inhibitor of apoptosis protein (IAP) family, which also includes cIAP1. It is the most potent caspase inhibitory IAP family member and a negative regulator of various apoptotic stimuli and death pathways.
XIAP overexpression in tumor cells is a well-described mediator of resistance to chemotherapy in many cancers and has been linked to tumor aggressiveness, making it an attractive target in cancer therapy where several therapeutic strategies have been investigated, such as SMAC mimetics.
Moreover, XIAP displays E3 ligase activity and leads to targeted proteins' ubiquitination and their subsequent degradation. This property can be exploited through a Proteolysis-targeting chimera (PROTAC) strategy. Therefore, new compounds targeting XIAP which exhibit dual roles: i) inhibition of XIAP anti-apoptotic function and ii) induction of targeted protein degradation, represent a promising therapeutic approach.
XIAP, like other IAP family proteins, contains BIR domains which interact with the IAP-binding motif of partners such as caspases. This interaction has been suggested to control pro- and anti-apoptotic activities.
XIAP compound characterization on the BIR2 and BIR3 domains enables accurate profiling and selectivity studies.
How it works
Assay principle
The HTRF XIAP BIR3 Binding Kit is a competitive assay format which uses XIAP BIR3 Red Ligand (based on A410099.1 derivative), a GST tagged human XIAP BIR3 binding domain, and an anti GST Terbium Cryptate-labeled antibody. XIAP binding compounds compete with the Red Ligand and thereby prevent FRET from occurring.

Assay protocol
The XIAP BIR3 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 XIAP BIR3 binding domain is then added, followed by the dispensing of the HTRF reagents: the anti GST antibody labeled with Terbium cryptate, and the ligand 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
Target Species-Human
Technology-TR-FRET
Therapeutic Area-Inflammation Neuroscience Oncology & Inflammation
Unit Size-10,000 assay points