Proteome Profiler Human XL Oncology Array ARY026: R&D Systems
Kit Summary

To detect differences in 84 cancer-related proteins between samples. No specialized equipment is necessary.

 

General Assay Principle

Carefully selected capture antibodies have been spotted in duplicate on nitrocellulose membranes. Cell culture supernates, cell lysates, tissue lysates, serum, plasma, urine, saliva, or human milk are diluted and incubated with the Human XL Oncology Array overnight. The array is washed to remove unbound proteins, followed by incubation with a cocktail of biotinylated detection antibodies. Streptavidin-HRP and chemiluminescent detection reagents are applied, and a signal is produced at each capture spot corresponding to the amount of protein bound. Chemiluminescence is detected in the same manner as a Western blot.

 

Kit Contents
  • 4 Array Membranes
  • 4-Well Multi-dish
  • Array Buffers
  • Wash Buffer
  • Detection Antibody Cocktail
  • Streptavidin-HRP
  • 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.

Stability and Storage

Reagents are stable for 12 months from date of receipt when stored in the dark at 2° C to 8°.

 

Simultaneously detect the levels of these cancer-related proteins in a single sample.
AFP ErbB4 MMP-2
Amphiregulin FGF basic MMP-3
Angiopoietin-1 FoxC2  MMP-9
ANGPTL4 FKHR MSP/MST1
ENPP-2/Autotaxin Galectin-3 MUC-1
AXL GM-CSF Nectin-4
BCL-X HCG Osteopontin
CA125/MUC-16 HGF R/c-Met p27/Kip1
E-Cadherin HIF-1alpha p53
VE-Cadherin HNF-3beta PDGF-AA
CAP-G HO-1/HMOX1 CD31/PECAM-1
CA-9 ICAM-1/CD54 Progesterone R
Cathepsin B CD25/IL-2 R alpha Progranulin
Cathepsin D IL-6 Prolactin
Cathepsin S CXCL8/IL-8 Prostasin
CEACAM-5 IL-18 Bpa E-Selectin
Decorin KLK-3/PSA Maspin
DKK-1 KLK-5 PAI-1/Serpin E1
DLL-1 KLK-6 SNAIL
EGF R/ErbB1 Leptin (OB) SPARC
Endoglin/CD105 Lumican Survivin
Endostatin CCL2/MCP-1 Tenascin-C
Enolase 2 CCL8/MCP-2 THBS-1
eNOS CCL7/MCP-3 TIE-2
EpCAM M-CSF UPA-1
ER-alpha Mesothelin VCAM-1
ErbB2 CCL3/MIP-1alpha VEGF
ErbB3 CCL20/MIP-3alpha Vimentin
Preparation and Storage
  • Stability & Storage
    Store the unopened product at 2 - 8 °C. Do not use past expiration date.
Related Research Areas
Assay Procedure

Refer to the product datasheet for complete product details.

Briefly, relative expression levels of human cancer-related proteins in samples can be determined using the following procedure:

  • Prepare membrane and incubate with prepared sample
  • Incubate the membrane with Detection Antibody Cocktail
  • Incubate the membrane array with Streptavidin-HRP
  • Develop the membrane array with Chemi Reagents 1 and 2
  • Expose the membrane array to autoradiography film
 
Kit Contents
  • Rectangular 4-Well Multi-dish
  • 4 Human XL Oncology Array nitrocellulose membranes spotted with 84 different antibodies to cancer-related proteins
  • Array Buffer 4
  • Array Buffer 6
  • Chemi Reagent 1
  • Chemi Reagent 2
  • Detection Antibody Cocktail, Human XL Oncology Array
  • Streptavidin-HRP
  • Transparency Overlay Template
  • Wash Buffer Concentrate (25X)

 

Other Supplies Required

Reagents

  • Pipettes and pipette tips
  • Gloves
  • 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)
  • Plastic wrap
  • Absorbent lab wipes (KimWipes® or equivalent)
  • Paper towels
  • X-ray film (Kodak BioMax Light-1) or equivalent
  • Flat-tipped tweezers

Equipment

  • Rocking platform shaker
  • Microcentrifuge
  • Autoradiography cassette
  • Film developer
  • 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 17
  • Aprotinin
  • Leupeptin (Catalog # EI002)
  • Pepstatin (Catalog # EI003)

Other Supplies Required for Tissue Lysate Samples

  • Protease Inhibitor Cocktail
  • Igepal® CA-630
  • Sodium deoxycholate
  • Sodium dodecyl sulfate

 

Procedure Overview

R&D Systems Protocol for Multiple Analyte Detection Using the Proteome Profiler Human XL Oncology Array Kit, Panel A (Catalog # ARY026)

Add 2 mL of Array Buffer 6 to each well of the supplied 4-Well Multi-dish.

ARY026 Step 1

Place each array membrane in a separate well of the 4-Well Multi-dish

Incubate for one hour on a rocking platform shaker.

ARY026 Step 2

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.

ARY026 Step 3

Replace the Array Buffer 6 in each well of the 4-Well Multi-dish with prepared samples.

Incubate overnight at 2 °C to 8 °C on a rocking platform.

Wash each array membrane 3 times with 1X Wash Buffer in a separate container.

Wash each well of the 4-Well Multi-dish with 1X Wash Buffer.

ARY026 Step 4

Add 30 µL of Detection Antibody Cocktail to 1.5 mL of Array Buffer 4/6 for each array.

ARY026 Step 5

Pipette 1.5 mL diluted Detection Antibody Cocktail into each well of the 4-Well Multi-dish.

ARY026 Step 6

Return each array membrane to the 4-Well Multi-dish containing diluted Detection Antibody Cocktail.

Incubate for one hour on a rocking platform shaker.

Wash each array membrane 3 times with 1X Wash Buffer in a separate container.

Wash each well of the 4-Well Multi-dish with 1X Wash Buffer.

ARY026 Step 7

Add 2 mL of diluted Streptavidin-HRP to each well of the 4-Well Multi-dish.

Place the array membrane in the diluted Streptavidin-HRP solution.

Incubate for 30 minutes on a rocking platform shaker.

Wash each array membrane 3 times with 1X Wash Buffer in a separate container.

ARY026 Step 8

Place the array membrane on a plastic sheet protector.

Pipette 1 mL of the prepared Chemi Reagent Mix evenly onto the membrane.

Cover the membrane with the top sheet of the plastic protector

Incubate for 1 minute.

Blot off excess Chemi Reagent Mix.

Wrap the membrane and sheet protector in plastic wrap.

Place the wrapped array membrane in an autoradiography film cassette and expose to X-ray film.

ARY026 Step 9
Citations:

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.

1 Citations: Showing 1 - 1

  1. CCL20 mediates RANK/RANKL-induced epithelial-mesenchymal transition in endometrial cancer cells
    Oncotarget, 2016;7(18):25328-39. 2016
Supplemental ELISA Products
Description Application Cat# Citations Images  

Lysis Buffer 17 (1 x 21 mL)

ELISA 895943 4
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