Human/Mouse/Rat Phospho-EGFR (Multi-site) Cell-Based ELISA (2)

RayBio® Cell-Based Human/Mouse/Rat EGFR (Multi-site) Phosphorylation ELISA Kit. Suitable for adherent whole cell lines.

CBEL-EGFR-SK
+

Antigen Information

Accession Number:
  • P00533
Gene ID:
  • 1956
Gene Symbols:
  • EGFR
  • ERBB
  • ERBB1
  • HER1
Protein Name & Synonyms:
Epidermal growth factor receptor (EC 2.7.10.1) (Proto-oncogene c-ErbB-1) (Receptor tyrosine-protein kinase erbB-1)

Assay Format

Pathway:
  • HER/ErbB Signaling
Specificity:
The antibodies provided in this kit recognizes human, mouse and rat EGFR phosphorylated at multiple sites (Tyrosine-phosphorylated EGFR, EGFR phosphorylated at site Tyrosine-1068, EGFR phosphorylated at site Tyrosine-845 and EGFR phosphorylated at site Tyrosine-992) and total EGFR for comparison.
Number of Targets Detected:
1
Species Detected:
  • Human
  • Mouse
  • Rat
Compatible Sample Types:
  • Adherent Cell Culture
Design Principle:
  • Cell-based
Method of Detection:
Colorimetric
Quantitative/Semi-Quantitative:
Semi-Quantitative
Solid Support:
96-well Microplate

Product Specifications

Size:
Two 96-Well Microplate Kit

Product Features

  • Site and signal pathway-specific
  • In vitro detection of adherent cell culture
  • No sample lysis needed
  • Compatible with a standard ELISA plate reader
  • Faster results than with ELISA
  • Adaptable for high-throughput screening and drug discovery

Application Notes

Kit Components
  • Cell Culture Microplate
  • Wash Buffer A
  • Wash Buffer B
  • Fixing Solution
  • HRP-Conjugated Secondary Antibody
  • Blocking Buffer
  • TMB One-Step Substrate
  • Stop Solution
  • Quenching Buffer
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
  1. Seed 10,000-30,000 cells into each well and incubate overnight.
  2. Apply various treatment, inhibitors or activators according to manufacture's instructions.
  3. Add 100 µl of Fixing Solution into each well and incubate for 20 min at RT with shaking.
  4. Add 200 µl of prepared 1X Quenching Buffer and incubate 20 min at RT.
  5. Add 200 µl of Blocking Solution and incubate for 1 h at 37 °C.
  6. Add 50 µl of 1X anti-phospho-protein specific antibody or anti-pan-protein specific antibody to each well and incubate for 2 h at RT.
  7. Add 50 µl of prepared 1X HRP-Anti-Rabbit or Mouse IgG and incubate for 1 h at RT.
  8. Add 100 µl of TMB One-Step Substrate Reagent to each well.
  9. Incubate 30 min at RT.
  10. Add 50 µl of Stop Solution to each well.
  11. Read at 450 nm immediately.

Storage/Stability

-20°C

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Usage

This product is furnished for LABORATORY RESEARCH USE ONLY.
Not for diagnostic or therapeutic use.