Revvity, 64RIPK1TPEG, HTRF Human Total RIPK1 Detection Kit, 500 Assay Points

Overview
The Total-RIPK1 r assay monitors total RIPK1, and can be used as a normalization assay with the Phospho-RIPK1 (Ser166) kit. t is compatible with the buffers from the phospho-RIPK1 the same lysate can be used for analyses of both the phosphorylated and the total protein populations. Receptor-Interacting serine/threonine-Protein Kinase 1, also called RIP1 or RIP) is a iator of apoptotic and necrotic cell death, as well as for inflammatory pathways eam of TNFR1 and other receptors. utophosphorylation at Serine 166 is considered to be a biomarker of the initiation of RIPK3/MLKL-dependent necroptosis. roptosis ogrammed necrotic cell death pathway, also called "inflammatory cell death", s closely associated with pathologies including inflammatory and neurodegenerative diseases, as well as cancer. as therefore become an important drug target in the pharmaceutical industry. Therapeutic strategies consist in developing specific small-molecule kinase inhibitors, such as the well-known Necrostatin-1 and Necrostatin-1s.
How it works
Total-RIPK1 assay principle
The HTRF Total-RIPK1 assay quantifies the expression level of RIPK1 in a cell lysate. Unlike Western Blot, the assay is entirely plate-based and does not require gels, electrophoresis, or transfer. The Total-RIPK1 assay uses two labeled antibodies: one coupled to a donor fluorophore, the other to an acceptor. Both antibodies are highly specific for a distinct epitope on the protein. In presence of RIPK1 in a cell extract, the addition of these conjugates brings the donor fluorophore into close proximity with the acceptor, and thereby generates a FRET signal. Its intensity is directly proportional to the concentration of the protein present in the sample, and provides a means of assessing the protein’s expression under a no-wash assay format.

 

Total-RIPK1 two-plate assay protocol
The two-plate protocol involves culturing cells in a 96-well plate before lysis, then transferring lysates to a 384-well low volume detection plate before the addition of Total-RIPK1 HTRF detection reagents. This protocol enables the cells' viability and confluence to be monitored.

 

Total-RIPK1 one-plate assay protocol
Detection of Total-RIPK1 with HTRF reagents can be performed in a single plate used for culturing, stimulation, and lysis. No washing steps are required. This HTS designed protocol enables miniaturization while maintaining robust HTRF quality. Application-Cell Signaling
Brand-HTRF
Detection Modality-HTRF
Lysis Buffer Compatibility-Lysis Buffer 1 Lysis Buffer 2 Lysis Buffer 3 Lysis Buffer 4 Lysis Buffer 5
Molecular Modification-Total
Product Group-Kit
Sample Volume-16 µL
Shipping Conditions-Shipped in Dry Ice
Target Class-Phosphoproteins
Target Species-Human
Technology-TR-FRET
Therapeutic Area-Inflammation Neuroscience Oncology & Inflammation
Unit Size-500 assay points