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Proteome Profiler Mouse XL Cytokine Array

Contains 4 membranes-each spotted in duplicate with 111 different cytokine antibodies
Detection of selected mouse cytokines in Balb/3T3 Cell Line Lysate
(3)
Detection of Selected Mouse Cytokines in CMT-93 Cell Line Lysate.
Detection of Selected Mouse Cytokines in Balb/3T3 Cell Line Supernate.
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ARY028
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Assay Procedure

Refer to the product datasheet for complete product details.

Briefly, relative expression levels of mouse cytokines and chemokines 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 Mouse XL Cytokine Array nitrocellulose membranes spotted with 111 different antibodies to mouse cytokines
  • Array Buffer 4
  • Array Buffer 6
  • Chemi Reagent 1
  • Chemi Reagent 2
  • Detection Antibody Cocktail, Mouse XL Cytokine Array
  • Streptavidin-HRP
  • Transparency Overlay Template
  • Wash Buffer Concentrate (25X)

 

Other Supplies Required

Reagents

Materials

  • 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 (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)
  • Triton X-100

 

Procedure Overview

R&D Systems Protocol for Multiple Analyte Detection Using the Proteome Profiler Mouse XL Cytokine Array Kit, Panel A (Catalog # ARY028)

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

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.

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

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.

Add 0.5 mL Array Buffer 4 to each sample.

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.

Add 0.5 mL Array Buffer 4 to each sample.
Add 30 µL of Detection Antibody Cocktail to 1.5 mL of Array Buffer 4/6 for each array.
Add 0.5 mL Array Buffer 4 to each sample.

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

Add 0.5 mL Array Buffer 4 to each sample.

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.

Add 0.5 mL Array Buffer 4 to each sample.

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.

Add 0.5 mL Array Buffer 4 to each sample.

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.

Add 0.5 mL Array Buffer 4 to each sample.

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Customer Reviews (20)

4.9 out of 5
20 customer ratings
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  • Proteome Profiler Mouse XL Cytokine Array
    Name: Anonymous
    Verified Customer | Posted 03/14/2021
    Proteome Profiler Mouse XL Cytokine Array Serum ARY028
  • Proteome Profiler Mouse XL Cytokine Array
    Name: Anonymous
    Verified Customer | Posted 09/17/2020
    Excellent antibody array to screen a multitude of cytokines, chemokines, and growth factors in one small strip of membrane. Readout is not a fluorescence so you don't need a special fluorescence reader. If you are doing Western blotting, you can do this easily.
    Proteome Profiler Mouse XL Cytokine Array Serum ARY028
  • Proteome Profiler Mouse XL Cytokine Array
    Name: Anonymous
    Verified Customer | Posted 02/12/2020
    Mouse breast tumor tissues
    Proteome Profiler Mouse XL Cytokine Array Serum ARY028
  • Proteome Profiler Mouse XL Cytokine Array
    Name: Anonymous
    Verified Customer | Posted 01/25/2020
  • Proteome Profiler Mouse XL Cytokine Array
    Name: Anonymous
    Verified Customer | Posted 01/06/2020
    Excellent array!!
    Proteome Profiler Mouse XL Cytokine Array Serum ARY028
  • Proteome Profiler Mouse XL Cytokine Array
    Name: Anonymous
    Verified Customer | Posted 06/06/2019
    Proteome Profiler Mouse XL Cytokine Array Serum ARY028
  • Proteome Profiler Mouse XL Cytokine Array
    Name: Anonymous
    Verified Customer | Posted 03/13/2019
  • Proteome Profiler Mouse XL Cytokine Array
    Name: Anonymous
    Verified Customer | Posted 12/05/2018
  • Proteome Profiler Mouse XL Cytokine Array
    Name: Anonymous
    Verified Customer | Posted 11/11/2018
    Excellent specificity and detection for large number of cytokines. Works perfectly with little optimization.
    Proteome Profiler Mouse XL Cytokine Array Serum ARY028
  • Proteome Profiler Mouse XL Cytokine Array
    Name: Anonymous
    Verified Customer | Posted 11/05/2018
    Pancreas lysate. 750ug protein/ membrane
    Proteome Profiler Mouse XL Cytokine Array Serum ARY028
  • Proteome Profiler Mouse XL Cytokine Array
    Name: Anonymous
    Verified Customer | Posted 07/18/2018
  • Proteome Profiler Mouse XL Cytokine Array
    Name: Anonymous
    Verified Customer | Posted 04/01/2018
  • Proteome Profiler Mouse XL Cytokine Array
    Name: Anonymous
    Verified Customer | Posted 03/22/2018
    Proteome Profiler Mouse XL Cytokine Array Serum ARY028
  • Proteome Profiler Mouse XL Cytokine Array
    Name: Anonymous
    Verified Customer | Posted 12/07/2017
    200 uL pooled serum from wild-type C57Bl/6 mice.5 min exposure time
    Proteome Profiler Mouse XL Cytokine Array Serum ARY028
  • Proteome Profiler Mouse XL Cytokine Array
    Name: Anonymous
    Verified Customer | Posted 08/17/2017
  • Proteome Profiler Mouse XL Cytokine Array
    Name: Anonymous
    Verified Customer | Posted 07/17/2017
    Proteome Profiler Mouse XL Cytokine Array Serum ARY028
  • Proteome Profiler Mouse XL Cytokine Array
    Name: Anonymous
    Verified Customer | Posted 05/23/2017
    Proteome Profiler Mouse XL Cytokine Array Serum ARY028
  • Proteome Profiler Mouse XL Cytokine Array
    Name: Anonymous
    Verified Customer | Posted 11/28/2016
    150 uL of mouse serum
    Proteome Profiler Mouse XL Cytokine Array Serum ARY028
  • Proteome Profiler Mouse XL Cytokine Array
    Name: Anonymous
    Verified Customer | Posted 09/30/2016
    This is a great kit. It provides excellent reliability and a large number of cytokines, chemokines to screening in a single assay. I strongly recommend it.
  • Proteome Profiler Mouse XL Cytokine Array
    Name: Anonymous
    Verified Customer | Posted 08/26/2016
    The sample was cell culture supernatant collected from MEF.
    Proteome Profiler Mouse XL Cytokine Array Serum ARY028

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FAQs

  • Will the Array identification number stamped on the Array membrane interfere with detection if it is not cut-off before the membrane is blocked?

    The dye used for printing the Array identification number on the membranes will fluoresce and interfere with the LI-COR detection. It is critical that the number is cut off before beginning the experiment. 

  • Could you tell us whether this MMP-9 antibody can detect TIMP and MMP-9 complex?

    We haven't performed a large amount of specificity testing per each spot in the arrays because these are qualitative screening tools. However, we would recommend treating the MMP-9 spot as a "Total MMP-9" detecting both free MMP-9 and MMP-9 in complex. If the MMP-9 spot turns out to be of high interest for the customer's study, they will want to examine this target in more detail with quantitative tools such as ELISA or Luminex.

Product Datasheets

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