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Human Phospho-CREB (S133) ELISA

Human Phospho-CREB (S133) ELISA

Manufacturer	Raybiotech
Category	Neuroscience Molecular Targets for Neuroscience Neural Signaling ELISA ELISA & Immunoassays ELISA Kits Immunoassay
Type	Elisa-Kit
Specific against	Human
Amount	5 Plate Kit
Item no.	PEL-CREB-S133-5
Targets	CREB1
eClass 6.1	32160605
eClass 9.0	32160605
Available
Species Detected	Human
Compatible Sample Types	Cell Lysates Tissue Lysates
Design Principle	Sandwich-based
Method of Detection	Colorimetric
Quantitative/Semi-Quantitative	Semi-Quantitative
Solid Support	96-well Microplate
Gene Symbols	CREB1
Kit Components	Pre- Coated 96-well Strip Microplate Wash Buffer Anti- Phospho Antibody H R P- Conjugated Secondary Antibody Assay Diluent T M B One- Step Substrate Stop Solution Lysis Buffer Positive Control Sample
Other Materials Required	Distilled or deionized water100 ml and 1 liter graduated cylinders Tubes to prepare sample dilutions Protease and Phosphatase inhibitors Precision pipettes to deliver 2 µl to 1 ml volumes Adjustable 1-25 ml pipettes for reagent preparation Benchtop rocker or shaker Microplate reader capable of measuring absorbance at 450 nm
Protocol Outline	Prepare all reagents and samples as instructed in the manual. Add 100 µl of sample or positive control to each well. Incubate 2.5 h at R T or O/ N at 4 ° C. Add 100 µl of prepared primary antibody to each well. Incubate 1 h at R T. Add 100 µl of prepared 1 X H R P- Streptavidin to each well. Incubate 1 h at R T. Add 100 µl of T M B One- Step Substrate Reagent to each well. Incubate 30 min at R T. Add 50 µl of Stop Solution to each well. Read at 450 nm immediately.
Protein Name & Synonyms	Cyclic AMP-responsive element-binding protein 1 (CREB-1) (cAMP-responsive element-binding protein 1)
Pathway	MAPK SignalingNeuroscience
Introduction	RayBio® Phospho-CREB (S133) ELISA kit is a very rapid, convenient and sensitive assay kit that can monitor the activation or function of important biological pathways in human cell lysates. By determining phosphorylated CREB protein in your experimental model system, you can verify pathway activation in your cell lysates. You can simultaneously measure numerous different cell lysates without spending excess time and effort in performing a Western Blotting analysis.This Sandwich ELISA kit is an in vitro enzyme-linked immunosorbent assay for the measurement of human phospho- CREB. An anti-pan CREB antibody has been coated onto a 96-well plate. Samples are pipetted into the wells and CREB present in a sample is bound to the wells by the immobilized antibody. The wells are washed and rabbit anti-CREB (S133) antibody is used to detect phosphorylated CREB. After washing away unbound antibody, HRP-conjugated anti-rabbit IgG is pipetted to the wells. The wells are again washed, a TMB substrate solution is added to the wells and color develops in proportion to the amount of CREB (S133) bound. The Stop Solution changes the color from blue to yellow, and the intensity of the color is measured at 450 nm.
Positive Control	HeLa cells were treated with PMA at 37°C for 20 min. Solubilize cells at 4 x 107 cells/ml in Cell Lysate Buffer. Serial dilutions of lysates were analyzed in this ELISA. Please see step 3 of Part VI Reagent Preparation for detail.
PMA Stimulation of HeLa Cell Lines	HeLa cells were treated or untreated with 250 nM PMA for 20 min. Cell lysates were analyzed using this phospho-ELISA and Western Blot.

Note: The presented information and documents (Manual, Product Datasheet, Safety Datasheet and Certificate of Analysis) correspond to our latest update and should serve for orientational purpose only. We do not guarantee the topicality. We would kindly ask you to make a request for specific requirements, if necessary.

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