Revvity, 64IDO1TPEG, HTRF Human Total IDO1 Detection Kit, 500 Assay Points

Overview
The Total IDO1 cell-based assay provides a convenient and accurate way of detecting Total IDO1 levels within cells.
IDO1 (Indoleamine 2,3 dioxygenase 1) is the most important rate-limiting enzyme of the kynurenine pathway due to its ability to catalyze the degradation of L-Tryptophan (Trp) to "kynurenine" metabolites (Kyn). Trp depletion and Kyn production promote immunosuppression by inhibiting CD8+ T effector cells and NK cells. It also increases the activity of CD4+ Treg cells and MDSCs (myeloid-derived suppressor cells). IDO1 also plays a key role in promoting new blood vessel development (angiogenesis).
IDO1 is overexpressed in different tumor cells and infiltrates immune cells, leading to tumor immune escape and neovascularization.
Although IDO1 inhibitors have been developed to inhibit tryptophan metabolism, these therapies have had limited success in prolonging patient survival. Recent efforts have shifted to TPD (targeted protein degradation) of IDO1 as a potentially more efficacious therapeutic strategy.
How it works
Total IDO1 assay principle

The Total IDO1 assay quantifies the expression level of IDO1 in a cell lysate. Unlike Western Blot, the assay is entirely plate-based, and does not require gels, electrophoresis, or transfer. The Total IDO1 assay uses two labeled antibodies, one coupled to a donor fluorophore and the other to an acceptor. Both antibodies are highly specific for a distinct epitope on the protein. In presence of IDO1 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 IDO1 two-plate assay protocol
The 2-plate protocol involves culturing cells in a 96-well plate before lysis, then transferring lysates into a 384-well low volume detection plate before the addition of the HTRF Total IDO1 detection reagents. This protocol enables the cells' viability and confluence to be monitored.

Total IDO1 one-plate assay protocol
Detection of Total IDO1 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
Molecular Modification-Total
Product Group-Kit
Sample Volume-16 µL
Shipping Conditions-Shipped in Dry Ice
Target Class-Phosphoproteins
Technology-TR-FRET
Unit Size-500 assay points