Phosphorylation ELISA Kits

Semi-Quantitative, Sandwich ELISAs for Detection of Phosphorylated Proteins in Cell Lysates

RayBio® Phosphorylation ELISA kits are rapid, convenient and sensitive assays that monitor the activation or function of important biological pathways in cell lysate samples. By determining phosphorylated protein in experimental model systems, researchers can verify pathway activation and simultaneously measure numerous different cell lysates without spending excess time and effort in performing Western Blot analyses.

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Phospho-ELISA kit


  • 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
  • High sensitivity
  • High quality
  • Cost-effective
  • Complete kit with pre-coated 96 well plate

Contents of Kit

  • 96 wells (12 strips x 8 wells) Coated With Antibody
  • Wash Buffer Concentrate (20x)
  • Positive Control
  • Assay Diluent
  • Detection Antibody
  • Secondary Antibody or HRP Streptavidin
  • TMB One-Step Substrate Reagent
  • Stop Solution
  • Cell Lysis Buffer

How It Works

Phospho-ELISA - How it works

This Sandwich ELISA kit is an in vitro enzyme-linked immunosorbent assay for the measurement of phosphorylated protein. An specific capture antibody (anti-pan-antibody or anti-phosphorylated antibody) has been coated onto a 96-well plate. Samples are pipetted into the wells and target protein present in a sample is bound to the wells by the immobilized antibody. The wells are washed and anti-phosphorylated antibody or anti-pan-antibody is used to detect target protein. After washing away unbound antibody, secondary antibody 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 target bound. The Stop Solution changes the color from blue to yellow, and the intensity of the color is measured at 450 nm.

GSK3b Positive Control

Representative Results

1. Recombinant Human EGF Stimulation of A431 Cell Lines:

A431 cells were treated or untreated with 100 ng/ml recombinant human EGF for 10 min. Cell lysates were analyzed using this phosphoELISA and Western Blot.

Phospho ELISA

2. Sensitivity:

The A431 cells were treated with 100 ng/mL recombinant human EGF for 20 minutes to induce phosphorylation of EGF R. Serial dilutions of lysates were analyzed in this ELISA and by Western blot. Immunoblots were incubated with anti-phospho-EGFR (Tyr 1086).

Phospho ELISA Sensitivity

Why RayBiotech ELISA Kits?

  • We manufacture, not resell, ELISA kits. Right here in the USA.
  • Our ELISAs are ISO certified and validated for exceptional reproducibility. Read more about our quality control processes.
  • All RayBio® ELISAs are backed by our 90-day 100% satisfaction guarantee.
  • Discounts and incentives available for publications citing our products, and for product reviews.


  1. Sok Kuan Wong, Kok-Yong Chin, Fairus Ahmad, Soelaiman Ima-Nirwana, Regulation of inflammatory response and oxidative stress by tocotrienol in a rat model of non-alcoholic fatty liver disease, Journal of Functional Foods, Volume 74, 2020, 104209, ISSN 1756-4646, [view publication]
    Product: Human/Mouse/Rat Phospho-NF-KB p65 (S536) ELISA
  2. Mohamed, Y.S., Abdelsalam, R.M., Attia, A.S. et al. Regulation of liver regeneration by prostaglandin E2 and thromboxane A2 following partial hepatectomy in rats. Naunyn-Schmiedeberg's Arch Pharmacol 393, 1437–1446 (2020). [view publication]
    Product: Human/Mouse/Rat Phospho-Stat 3 (Y705) ELISA
  3. Namgyal D, Chandan K, Sultan A, Aftab M, Ali S, Mehta R, El-Serehy HA, Al-Misned FA, Sarwat M. Dim Light at Night Induced Neurodegeneration and Ameliorative Effect of Curcumin. Cells. 2020 Sep 13;9(9):2093. doi: 10.3390/cells9092093. PMID: 32933226; PMCID: PMC7565558. [view publication]
    Product: Human Phospho-CREB (S133) ELISA
  4. Hegazy N, Rezq S, Fahmy A. Mechanisms Involved in Superiority of Angiotensin Receptor Blockade over ACE Inhibition in Attenuating Neuropathic Pain Induced in Rats. Neurotherapeutics. 2020 Jul;17(3):1031-1047. doi: 10.1007/s13311-020-00912-8. PMID: 32804335; PMCID: PMC7609714. [view publication]
    Product: Human/Mouse/Rat Phospho-Stat 3 (Y705) and Total Stat 3 ELISA
  5. Sugimoto A, Kataoka TR, Ito H, Kitamura K, Saito N, Hirata M, Ueshima C, Takei Y, Moriyoshi K, Otsuka Y, Nishikori M, Takaori-Kondo A, Haga H. SLAM family member 8 is expressed in and enhances the growth of anaplastic large cell lymphoma. Sci Rep. 2020 Feb 13;10(1):2505. doi: 10.1038/s41598-020-59530-1. PMID: 32054954; PMCID: PMC7018816. [view publication]
    Product: Human/Mouse/Rat Phospho-SHP2 (Y542) and Total SHP2 ELISA

More references are listed per product on the individual product pages.