Proteome Profiler Human Phospho-Immunoreceptor Array Kit
Proteome Profiler Human Phospho-Immunoreceptor Array Kit Summary
A membrane-based antibody array for the parallel determination of the relative phosphorylation of human immunoreceptors. Validated for analyte detection in cell lysates.
- Detects phosphorylation of 59 human ITAM/ITIM-associated immunoreceptors simultaneously
- Requires no specialized equipment
Principle of the Assay
The Proteome Profiler Human Phospho-Immunoreceptor Array Kit is a membrane-based sandwich immunoassay. Capture antibodies spotted in duplicate on nitrocellulose membranes bind to specific target proteins present in the sample (Step 1). Tyrosine phosphorylation of the captured proteins is detected with an HRP-conjugated pan phospho-tyrosine antibody (Step 2) and then visualized using chemiluminescent detection reagents (Step 3). The signal produced is proportional to the amount phosphorylation in the bound analyte.
Why Use an Antibody Array to Detect Receptor Phosphorylation?
Determining the phosphorylation of multiple immunoreceptors in a single sample can be expensive, time consuming and can require specialized equipment. Performing multiple immunoprecipitations and Western blots requires time, labor, and reagents. The use of a multiplex antibody array to detect multiple phosphorylations in a single sample can be cost-effective and also save time and sample.
This kit contains the following azide-free, fluorochrome-conjugated antibodies to verify human stem cell pluripotency:
- 4 Array Membranes
- 4-Well Multi-dish
- Array Buffers
- Lysis Buffer
- Wash Buffer
- HRP-conjugated Anti-phospho-tyrosine Detection Antibody
- Chemiluminescent Detection Reagents
- Transparency Overlay Template
- Detailed Protocol
For a complete list of the kit contents and necessary materials, please see the Materials Provided/Other Supplies Required sections of the product datasheet.
|Simultaneously detect the relative phosphorylation levels of these immunoreceptors in a single sample.|
|CEACAM-1||Integrin beta 3/CD61||Siglec-3/CD33|
|Fc epsilon RII/CD23||MDL-1/CLEC5A||TREM-2|
|Fc gamma RIIA||NKp30/NCR3||TREML1/TLT-1|
|Fc gamma RIIIA/B||NKp44/NCR2|
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Figure 1A: The Human Phospho-Immunoreceptor Array detects multiple tyrosine phosphorylated immunoreceptors in lysates prepared from cells activated by antibodymediated cross-linking of cell-surface immunoreceptors. THP-1 human acute monocytic leukemia cells were incubated with goat anti-Fcγ RIIA antibody (R&D Systems, Catalog # AF1875) or normal goat IgG (R&D Systems, Catalog # AB-108-C) followed by incubation with a donkey anti-goat IgG antibody (R&D Systems, Catalog # AF109) for 5 minutes [Maresco, D.L. et al. (1999) J. Immunol. 162:6458].
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Figure 1B: Jurkat human acute T cell leukemia cells were incubated with a mouse monoclonal anti-CD3ε antibody (R&D Systems, Catalog # MAB100) or mouse IgG1 isotype control (R&D Systems, Catalog # MAB002) followed by incubation with a goat anti-mouse antibody (R&D Systems, Catalog # AF007) for 5 minutes. 100 µg of lysate was run on each array.
R&D Systems Human Phospho-Immunoreceptor Array (Catalog # ARY004) is a rapid, sensitive, and economical tool used to simultaneously detect the relative phosphorylation levels of 59 ITAM/ITIM-associated immunoreceptors, avoiding numerous immunoprecipitations and/or Western blots.
Citations for Proteome Profiler Human Phospho-Immunoreceptor Array Kit
R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.
Citations: Showing 1 - 4
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Molecular basis of reduced LAIR1 expression in childhood severe malarial anaemia: Implications for leukocyte inhibitory signalling
Authors: AO Achieng, B Guyah, Q Cheng, JM Ong'echa, C Ouma, CG Lambert, DJ Perkins
EBioMedicine, 2019;45(0):278-289. 2019
Evidence for C1q-mediated crosslinking of CD33/LAIR-1 inhibitory immunoreceptors and biological control of CD33/LAIR-1 expression
Authors: M Son, B Diamond, BT Volpe, CB Aranow, MC Mackay, F Santiago-S
Sci Rep, 2017;7(1):270. 2017
Fibroblast Growth Factor Receptor 3 (FGFR3) Associated with the CD20 Antigen Regulates the Rituximab-induced Proliferation Inhibition in B-cell Lymphoma Cells.
Authors: Kotani N, Ishiura Y, Yamashita R, Ohnishi T, Honke K
J Biol Chem, 2012;287(44):37109-18. 2012
Surface molecule CD229 as a novel target for the diagnosis and treatment of multiple myeloma.
Authors: Atanackovic D, Panse J, Hildebrandt Y, Jadczak A, Kobold S, Cao Y, Templin J, Meyer S, Reinhard H, Bartels K, Lajmi N, Zander AR, Marx AH, Luetkens T, Bokemeyer C, Kroger N
Haematologica, 2011;0(0):. 2011
The insert states to lyse 1 x 107 cells in 1 mL Lysis Buffer 17. What is the approximate protein concentration of the resulting lysate?
Lysing following recommendations in the product insert produces a lysate with a protein concentration of about 1.5-2.0 mg/mL. Quantitation of sample protein concentration using a total protein assay is recommended.
Do I need to use the protease inhibitors (e.g. aprotinin, Ipegal) if I only plan to use cell supernatants?
The proteinase inhibitors are recommended for cell and tissue lysates only. You do not need them for cell culture supernates.
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