The RayBiotech Human Phospho-Glucocorticoid Receptor (S226) ELISA Kit measures phosphorylated Glucocorticoid Receptor (Ser226) and total (pan) Glucocorticoid Receptor in cell and tissue lysates.
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Product Description
Specifications
| Size | 1 Plate Kit, 2 Plate Kit, 5 Plate Kit |
| Estimated Lead Time | 1-2 business days |
| Species | Human |
| Specificity | The antibody pair provided in this kit recognizes Human Glucocorticoid Receptor phosphorylated at site Serine-226. |
| Compatible Sample Types | Tissue Lysates, Cell Lysates |
| Accession Number | P04150 |
| Gene Symbols | NR3C1, GRL |
| Protein Name / Synonyms | Glucocorticoid receptor (GR) (Nuclear receptor subfamily 3 group C member 1) |
| Quantitative/Semi-Quantitative | Semi-Quantitative |
| Solid Support | 96-well Microplate |
| Design Principle | Sandwich-based |
| Method Of Detection | Colorimetric |
| Research Area | Post-Translational Modifications, Inflammation, Phosphorylation, Glucocorticoid Receptor |
| Shipping Type | Blue ice |
| Storage | -20°C |
Product Features
- Simultaneously measure phosphorylated protein and pan protein in one experiment (for normalization purpose)
- Screen numerous different cell lysates without performing a Western Blot analysis
- Minimal hands-on time, convenient, and non-radioactive material
Application Notes
Kit Components
- Pre-Coated 96-well Strip Microplate
- Wash Buffer
- Anti-Phospho Antibody
- Anti-Pan Antibody (*not included in phospho-only version)
- HRP-Conjugated Secondary Antibody
- Assay Diluent
- TMB One-Step Substrate
- Stop Solution
- Lysis Buffer
- Positive Control Sample
Other Materials Required
- Distilled or deionized water
- 100 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 RT or O/N at 4 °C.
- Add 100 µl of prepared primary antibody to each well.
- Incubate 1 h at RT.
- Add 100 µl of prepared 1X HRP-Streptavidin to each well.
- Incubate 1 h at RT.
- Add 100 µl of TMB One-Step Substrate Reagent to each well.
- Incubate 30 min at RT.
- Add 50 µl of Stop Solution to each well.
- Read at 450 nm immediately.
Typical Data
Positive Control
A431 cells were treated with recombinant human EGF at 37°C for 20 min. Cells were solubilzed at 4 x 107 cells/ml in lysis buffer. Serial dilutions of lysates were analyzed in this ELISA. Please see step 3 of Part VI. Reagent Preparation for details.
Calyculin A and PMA Stimulation of A549 Cell Line
A549 cells were treated with Calyculin A and PMA. Cell lysates were analyzed using this phosphoELISA and Western Blot.
Storage/Stability
Upon receipt, the kit should be stored at €“20°C. Please use within 6 months from the date of shipment.