図1。Treatment of ACHN cells with Human Interleukin-4 (hIL-4) stimulates phosphorylation of Stat6 at Tyr641 but does not effect the level of total Stat6. The relationship between lysate protein concentration from untreated and hIL-4-treated ACHN cells and the absorbance at 450 nm using the FastScan™ Phospho-Stat6 (Tyr641) ELISA Kit #58967 is shown in the upper figure. The corresponding western blots using phospho-Stat6 (Tyr641) antibody (left panel) and Stat6 antibody (right panel) are shown in the lower figure. After serum starvation, ACHN cells were treated with 100 ng/ml hIL-4 #8919 for 15 minutes at 37°C and then lysed.
|Product Includes||Volume (with Count)||Solution Color|
|FastScan™ ELISA Microwell Strip Plate, 96 Well 53257||1 x 96 tests|
|Stat6 Rabbit Capture mAb||1 x 1 ea||Green (Lyophilized)|
|Phospho-Stat6 (Tyr641) Rabbit HRP-linked mAb||1 x 1 ea||Red (Lyophilized)|
|FastScan™ ELISA Capture Antibody Diluent||1 x 3 ml||緑|
|FastScan™ ELISA HRP Antibody Diluent||1 x 3 ml|
|TMB Substrate 7004||1 x 11 ml|
|STOP Solution 7002||1 x 11 ml|
|Sealing Tape||1 x 1 ea|
|ELISA Wash Buffer (20X) 9801||1 x 25 ml|
|FastScan™ ELISA Cell Extraction Buffer (5X) 69905||1 x 10 ml|
|FastScan™ ELISA Cell Extraction Enhancer Solution (50X) 25243||1 x 1 ml|
|FastScan™ ELISA Kit #58967 Positive Control Type 1||1 x 1 ea|
NOTE: Prepare solutions with deionized/purified water or equivalent.
*IMPORTANT: The provided FastScan™ ELISA Cell Extraction Enhancer Solution (50X) may precipitate when stored at 4°C. この場合は37°Cで短時間温め、穏やかに攪拌して溶解してください。FastScan™ ELISA Cell Extraction Enhancer Solution (50X) は、沈殿を防ぐために室温で保管することができます。
NOTE: The 1X Cell Extraction Buffer contains phosphatase inhibitors. 細胞溶解の直前に、プロテアーゼ阻害剤を1X Cell Extraction Bufferに追加してください。追加のホスファターゼ阻害剤を加えることもできます (この場合はProtease/Phosphatase Inhibitor Cocktail (100X) #5872などを別途お求めください)。
NOTE: A select number of FastScan™ ELISA kits do not contain a positive control, please refer to Product Includes table on the datasheet for a list of included reagents. Should you need support on how to generate a positive control for those kits, contact CST technical support at firstname.lastname@example.org.
NOTE: Equilibrate all materials and prepared reagents to room temperature prior to running the assay.
*NOTE: Certain FastScan™ ELISA Kits may require additional washes at this step. 追加の洗いが必要な場合は、キットのデータシートにある製品の「説明」欄に明記してあります。
Protocol Id: 1924
The FastScan™ Phospho-Stat6 (Tyr641) ELISA Kit is a sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of Stat6 when phosphorylated at Tyr641. To perform the assay, sample is incubated with a capture antibody conjugated with a proprietary tag and a second detection antibody linked to HRP, forming a sandwich with phospho-Stat6 (Tyr641) in solution. This entire complex is immobilized to the plate via an anti-tag antibody. The wells are then washed to remove unbound material. TMB is then added. The magnitude of observed signal is proportional to the quantity of phospho-Stat6 (Tyr641). Antibodies in kit are custom formulations specific to kit.
The FastScan™ Phospho-Stat6 (Tyr641) ELISA Kit detects endogenous levels of Stat6 when phosphorylated at Tyr641 as shown in Figure 1. This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species.Species Reactivity:
Upon activation by Janus kinases, Stat6 translocates to the nucleus where it regulates cytokine-induced gene expression. Stat6 is activated via phosphorylation at Tyr641 and is required for responsiveness to IL-4 and IL-13 (1-4). In addition, Stat6 is activated by IFN-α in B cells, where it forms transcriptionally active complexes with Stat2 and p48 (5,6). Protein phosphatase 2A is also involved in regulation of IL-4-mediated Stat6 signaling (7).
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Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
FastScan™ ELISA is a trademark of Cell Signaling Technology, Inc.
U.S. Patents 9,086,407, 9,261,500, and 9,476,874, foreign equivalents, and child patents deriving therefrom.