Carefully selected capture antibodies have been spotted in duplicate on nitrocellulose membranes. Cell culture supernatant, cell lysate, or serum samples are diluted and mixed with a cocktail of biotinylated detection antibodies. The sample/antibody mixture is then incubated with the array. Any cytokine/detection antibody complex present is bound by its cognate immobilized capture antibody on the membrane. Streptavidin-Horseradish Peroxidase and chemiluminescent detection reagents are added, and a signal is produced in proportion to the amount of cytokine bound. Chemiluminescence is detected in the same manner as a Western blot.
2 Hematopoeitic Array and Membranes
2 Common Analyte Array Membranes
Detection Antibody Cocktail
Chemiluminescent Detection Reagents
Transparency Overlay Template
For a complete list of the kit contents and necessary materials, please see the Materials Provided and Other Materials Required sections of the product datasheet.
Simultaneously detect the levels of these soluble receptors in a single sample.
Plastic container with the capacity to hold 50 mL (for washing the arrays)
Plastic transparent sheet protector (trimmed to 10 cm x 12 cm and open on three sides)
Absorbent lab wipes (KimWipes® or equivalent)
X-ray film (Kodak BioMax™ Light-1) or equivalent
Rocking platform shaker
Flatbed scanner with transparency adapter capable of transmission mode
Computer capable of running image analysis software and Microsoft Excel
Other Supplies Required for Cell Lysate Samples
Phosphate-Buffered Saline (PBS)
Lysis buffer (1% Igepal CA-630, 20 mM Tris-HCl (pH 8.0), 137 mM NaCl, 10% glycerol, 2 mM EDTA, 10 µg/mL Aprotinin, 10 µg/mL Leupeptin, and 10 µg/mL Pepstatin)
Other Supplies Required for Tissue Lysate Samples
PBS with protease inhibitors (10 µg/mL Aprotinin, 10 µg/mL Leupeptin, and 10 µg/mL Pepstatin)
Add 2 mL of Array Buffer 6 to each well of the supplied 4-Well Multi-dish.
Place each array membrane in a separate well of the 4-Well Multi-dish.
Incubate for one hour on a rocking platform shaker.
Add 0.5 mL Array Buffer 4 to each sample.
Adjust volume of each sample to final volume of 1.5 mL with Array Buffer 6.
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.
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