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Email. hkkim@jcbio.co.kr

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Phospho-STAT1 (Tyr701) Cellular Assay Kit HTRF® cell-based immunoassay for quantification of phospho-STAT1 in JAK/STAT signaling Cisbio's cell-based homogeneous HTRF® phospho-STAT1 assay (Tyr701), is designed for the quantitative detection of STAT1 modulation, phosphorylated on tyrosine 701. Stimulated by growth factors and chemokines, like IFN-γ induced JAK/STAT signaling, STAT1 acts as an important transcriptional activator. Thus the assay serves as a readout for JAK inhibitors in oncology, infectiology and inflammation. This simple add-and-read assay is ideal for screening small molecules, compounds or biologics directly in cells. The phospho-STAT1 ready-to-use kit contains all the reagents you need and offers enhanced convenience over other immunoassay technologies, being easily amenable to high throughput screening and automated platforms.    Features Cell-based immunoassay as IFN-γ induced JAK readout High assay performance and robust results Truly homogeneous protocol with no wash steps More convenient and faster than ELISA or WB Low sample volume required and simple protocol   Applications Cellular detection and quantification of phospho-STAT1 (Tyr701)   Functional screening of activating receptor compounds in JAK1/STAT1 pathway   Oncology, infectiology, inflammation   Assay Principle HTRF® - the homogeneous cell-based sandwich immunoassay    The phospho-STAT1 assay is based on a TR-FRET sandwich immunoassay format comprising two specific anti-STAT1 antibodies, one labeled with a cryptate as donor and the other with d2 as acceptor. The phospho-STAT1 antibodies bind the phosphorylated residue, and the proximity of donor and acceptor will then lead to a fluorescent TR-FRET signal. The signal intensity is proportional to the substrate phosphorylation. The protocol is optimized for a 384-well plate format, but can easily be further miniaturized or upscaled. Only low sample volumes are needed. The detection reagents may be pre-mixed and added in a single dispensing step for direct detection. No washing is needed at any step.   The phospho-STAT1 assay kit can be run with frozen cell lysates or fresh cells in culture. After cell lysis, phospho-STAT1 can be quantitatively detected using the HTRF phospho-STAT1 kit reagents and most TR-FRET multimode plate readers.     Two-plate assay protocol For added flexibility, the assay can be run under a two-plate assay protocol, where cells are plated and treated in a 96-well culture plate. For the detection of phosphorylated STAT1, lysates are subsequently transferred to a 384 small volume assay plate where the HTRF reagents are added. This also enables the monitoring of cell viability and confluence in an appropriate cell culture plate.    

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