Human Phospho-Axl DuoSet IC ELISA
R&D Systems, part of Bio-Techne | Catalog # DYC2228-2
Key Product Details
Assay Type
Solid Phase Sandwich ELISA
Sample Type
Cell Lysates
Reactivity
Human
Human Phospho-Axl DuoSet IC ELISA Features
- Optimized capture and detection antibody pairings and recommended concentrations save lengthy development time
- Development protocols are provided to guide further assay optimization
- Assay can be customized to your specific needs
- Available in 2, 5, and 15-(96-well) plate pack sizes
- Economical alternative to Western blot
Product Summary for Human Phospho-Axl DuoSet IC ELISA
This DuoSet IC ELISA contains the basic components required for the development of sandwich ELISAs to measure tyrosine-phosphorylated human Axl in cell lysates. An immobilized capture antibody specific for Axl binds both phosphorylated and unphosphorylated Axl . After washing away unbound material, an HRP-conjugated detection antibody is used to detect only phosphorylated receptor, utilizing a standard HRP format.
Product Specifications
Assay Format
96-well strip plate (sold separately)
Sample Volume Required
Cell lysates (100 µL)
Detection Method
Colorimetric ELISA - 450nm (TMB)
Conjugate
Biotin
Specificity
Label
HRP
Scientific Data Images for Human Phospho-Axl DuoSet IC ELISA
Figure 1. The Human Phospho-Axl DuoSet IC ELISA is more sensitive than immunoprecipitation (IP)-Western analysis
Human glioblastoma cell line, A172, was treated with conditioned media from the human lung cell line, WI38, for fifteen minutes to induce tyrosine phosphorylation of Axl. Serial dilutions of lysates were analyzed by (A) IP-Western blot and (B) this ELISA . IPs were done using an anti-Axl monoclonal antibody and anti-mouse IgG agarose. Immunoblots were incubated with a biotinylated anti-phosphotyrosine monoclonal antibody (Catalog # BAM1676) to detect phospho-Axl (p-Axl). Bands were visualized with Streptavidin-HRP (Cat # DY998) followed by chemiluminescent detection.
Figure 2. The Human Phospho-AxlDuoSet IC ELISA detects ligand-induced Axl tyrosine phosphorylation
A172 cells were untreated or treated with conditioned media from the human lung cell line, WI38, for fifteen minutes. ELISA and IP-Western blot (inset) analyses were done using 100 μg and 400 μg of lysate, respectively. IP-Western blots for phospho-Axl (p-Axl) were done as described in Figure 1. Blots were stripped and total Axl was detected using a biotinylated anti-Axl polyclonal antibody (Catalog # BAF154).
Figure 3. The specificity of the Human Phospho-AxlDuoSet IC ELISA is confirmed by receptor competition
A172 cells were treated with conditioned media from the human lung cell line WI38 for fifteen minutes. The indicated amounts of recombinant extracellular domains of human Axl (Catalog # 154-AL), human Dtk (Catalog # 859-DK) or human Trk A (Catalog # 175-TK) were added to 125 μg lysate and analyzed using this ELISA. Competition was observed only with recombinant Axl.Kit Contents for Human Phospho-Axl DuoSet IC ELISA
- Capture Antibody
- Conjugated Detection Antibody
- Calibrated Immunoassay Standard or Control
Other Reagents Required
PBS: (Catalog # DY006), or 137 mM NaCl, 2.7 mM KCl, 8.1 mM Na2HPO4, 1.5 mM KH2O4, pH 7.2 - 7.4, 0.2 µm filtered
Wash Buffer: (Catalog # WA126), or equivalent
Lysis Buffer*
IC Diluent*
Blocking Buffer*
Substrate Solution: 1:1 mixture of Color Reagent A (H2O2) and Color Reagent B (Tetramethylbenzidine) (Catalog # DY999)
Stop Solution: 2 N H2SO4 (Catalog # DY994)
Microplates: R&D Systems (Catalog # DY990), or equivalent
Plate Sealers: ELISA Plate Sealers (Catalog # DY992), or equivalent
*For the Lysis Buffer, IC Diluent, and Blocking BUffer recommended for a specific DuoSet ELISA Development Kit, please see the product
Preparation and Storage
Shipping
The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage
Store the unopened product at 2 - 8 °C. Do not use past expiration date.
Background: Axl
Long Name
Axl Receptor Tyrosine Kinase
Alternate Names
AI323647, ARK, JTK11, Tyro7, UFO
Gene Symbol
AXL
Additional Axl Products
Product Documents for Human Phospho-Axl DuoSet IC ELISA
Product Specific Notices for Human Phospho-Axl DuoSet IC ELISA
For research use only
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