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    membrane-fraction-wb-cocktail-ab140365.pdf

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Kits/ Lysates/ Other Kits Antibody Cocktails Membrane
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Membrane Fraction WB Cocktail (ab140365)

  • Datasheet
  • SDS
Submit a review Q&A (2)References (4)

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Membrane Fraction Western Blot Cocktail - Component Seperation
  • Membrane Fraction Western Blot Cocktail - Specificity
  • Membrane Fraction Western Blot Cocktail - Specificity
  • Membrane Fraction Western Blot Cocktail - Cross Reactivity

Key features and details

  • Sample type: Cell culture extracts, Cell Lysate, Tissue Extracts

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Primary
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Anti-SIRT2 antibody [EPR20411-105] (ab211033)
Preparation
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Plasma Membrane Protein Extraction Kit (ab65400)

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Overview

  • Product name

    Membrane Fraction WB Cocktail
  • Sample type

    Cell culture extracts, Tissue Extracts, Cell Lysate
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Product overview

    ab140365 contains 5 mAbs each targeting proteins located in different compartments of the cell. The presence of plasma membrane is shown by Anti-Sodium Potassium ATPase; endoplasmic reticulum is detected by GRP78; mitochondrial by ATP5A; cytosol by Anti-GAPDH; and nucleus by Anti-Histone H3 (di methyl K9). This cocktail is suitable for detecting the purity of cellular fractions using kit ab139409.

  • Tested applications

    Suitable for: WBmore details

Properties

  • Storage instructions

    Store at -20°C. Please refer to protocols.
  • Components 200 µl
    2500X HRP Conjugated Secondary Antibody Cocktail 1 x 60µl
    250X Membrane Fraction Western Blot Cocktail 1 x 200µl
  • Research areas

    • Kits/ Lysates/ Other
    • Kits
    • Antibody Cocktails
    • Membrane

Associated products

  • Related Products

    • Organelle Detection Western Blot Cocktail (ab133989)
    • Endoplasmic Reticulum Fraction Western Blot Cocktail (ab139415)
    • Mitochondria Fraction Western Blot Cocktail (ab139416)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab140365 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
WB
Use at an assay dependent concentration.

Primary antibody cocktail = 1/250 dilution

Secondary antibody cocktail = 1/2500 dilution

Notes
WB
Use at an assay dependent concentration.

Primary antibody cocktail = 1/250 dilution

Secondary antibody cocktail = 1/2500 dilution

Images

  • Membrane Fraction Western Blot Cocktail - Component Seperation
    Membrane Fraction Western Blot Cocktail - Component Seperation
    Performed under reducing conditions. All blocking and antibody incubation steps were done in 5% milk in TBST. Developed using the ECL technique. Exposure time: 5 mins.

    Sample 1: Marker
    Samples 2-5: Hela Cell Lysate – 20 µg

    Primary:
    Lane 1: none
    Lane 2: Anti- Sodium Potassium ATPase antibody – Plasma Membrane Marker
    Lane 3: Anti-GRP78 antibody – Endoplasmic Reticulum Marker
    Lane 4: Anti- ATP5a antibody – Mitochondrial Membrane Marker
    Lane 5: Anti- GAPDH antibody – Cytosolic Marker
    Lane 6: Anti- Histone H3 (di methyl K9) antibody – Nuclear Marker
    Lane 7: Assembled Membrane Antibody Cocktail

    Secondary:HRP conjugated secondary antibody cocktail

    Predicted/observed band sizes:
    Sodium Potassium ATPase = 112 kDa/100 kDa
    GRP78 band size: 78 kDa / 75 kDa
    ATP5A band size: 60 kDa / 60 kDa
    GAPDH band size: 37 kDa / 38 kDa
    Histone H3 (di methyl K9) band size: 17 kDa / 17 kDa
  • Membrane Fraction Western Blot Cocktail - Specificity
    Membrane Fraction Western Blot Cocktail - Specificity

    Developed by IR scanning. HeLa cell lysate prepared using Kit ab139409. Exposure time: 5 mins . Samples run on a 4-20% gradient acrylamide gel. All blocking and antibody incubation steps were done in 5% milk, 20mM Tris-HCl, 0.1% TWEEN-20
    Lane 1 : HeLa Whole Cell Lysate
    Lane 2 : HeLa Cytosolic Fraction Lysate
    Lane 3 : HeLa Membrane Fraction Lysate
    Lane 4 : HeLa Nuclear Fraction Lysate
    Primary antibody: Membrane Fraction Western Blot cocktail
    Secondary: IR800 GAM (1/10000) and IR690 GAR (1/10000)
    Predicted/Observed band sizes:
    Sodium Potassium ATPase: 112 kDa/100 kDa
    GRP78: 78 kDa/75 kDa
    ATP5A: 60 kDa/60 kDa
    GAPDH: 37 kDa/37kDa
    Histone H3 (di methyl K9): 17 kDa/17kDa
    Notes: Percentage of antibody signal (Y axis) represents the signal present in individual fractions as a proportion of the sum of all three fractions (cytoplasmic, membrane and nuclear)

  • Membrane Fraction Western Blot Cocktail - Specificity
    Membrane Fraction Western Blot Cocktail - Specificity
    Developed using ECL technique under reducing conditions. HeLa lysate prepared using the Membrane Fractionation Kit (ab139409). Exposure time: 5 mins. on a 4-20% gradient acrylamide gel, blocking and antibody incubation steps done in 5% milk, 20mM Tris-HCl, 0.1% TWEEN-20
    Lane 1: Marker
    Lane 2 : HeLa Whole Cell Lysate
    Lane 3 : HeLa Cytosolic Fraction Lysate
    Lane 4 : HeLa Membrane Fraction Lysate
    Lane 5 : HeLa Nuclear Fraction Lysate
    Primary antibody: Membrane Fraction Western Blot cocktail
    Secondary: HRP conjugated secondary antibody cocktail
    Predicted/Observed band sizes:
    Sodium Potassium ATPase: 112 kDa/100 kDa
    GRP78: 78 kDa/75 kDa
    ATP5A: 60 kDa/60 kDa
    GAPDH: 37 kDa/37kDa
    Histone H3 (di methyl K9): 17 kDa/17kDa
    Notes: Percentage of antibody signal represents signal present in individual fractions as a proportion of the sum of all three fractions (cytoplasmic, membrane and nuclear)
  • Membrane Fraction Western Blot Cocktail - Cross Reactivity
    Membrane Fraction Western Blot Cocktail - Cross Reactivity
    Samples run on a 4-20% gradient acrylamide gel. Performed under reducing conditions. All blocking and antibody incubation steps were done in 5% milk, 20mM Tris-HCl, 0.1% TWEEN-20. Developed using the ECL technique. Exposure time: 5 mins.

    Lane 1: Marker
    Lane 2: Human heart homogenate Lysate – 20 µg
    Lane 3: Hela Cell Lysate – 20 µg
    Lane 4: Mouse heart homogenate Lysate - 20 µg
    Lane 5: NIH3T3 Cell Lysate – 20 µg
    Lane 6: Rat heart homogenate Lysate – 20 µg
    Lane 7: H9C2 Cell Lysate – 20 µg

    Primary antibody: Membrane Fraction Western blot cocktail at 1/250 dilution

    Secondary antibody: HRP conjugated secondary antibody cocktail at 1/2500 dilution

    Predicted/observed band sizes:
    Sodium Potassium ATPase = 112 kDa/100 kDa
    GRP78 band size: 78 kDa / 75 kDa
    ATP5A band size: 60 kDa / 60 kDa
    GAPDH band size: 37 kDa / 38 kDa
    Histone H3 (di methyl K9) band size: 17 kDa / 17 kDa

Protocols

To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.

Click here to view the general protocols

Datasheets and documents

  • SDS download

  • Datasheet download

    Download

References (4)

Publishing research using ab140365? Please let us know so that we can cite the reference in this datasheet.

ab140365 has been referenced in 4 publications.

  • Buzhdygan TP  et al. The psychoactive drug of abuse mephedrone differentially disrupts blood-brain barrier properties. J Neuroinflammation 18:63 (2021). PubMed: 33648543
  • Otto M  et al. 12(S)-HETE mediates diabetes-induced endothelial dysfunction by activating intracellular endothelial cell TRPV1. J Clin Invest 130:4999-5010 (2020). PubMed: 32584793
  • Foryst-Ludwig A  et al. Adipose Tissue Lipolysis Promotes Exercise-induced Cardiac Hypertrophy Involving the Lipokine C16:1n7-Palmitoleate. J Biol Chem 290:23603-15 (2015). WB ; Mouse . PubMed: 26260790
  • Bohuslavova R  et al. Partial deficiency of HIF-1a stimulates pathological cardiac changes in streptozotocin-induced diabetic mice. BMC Endocr Disord 14:11 (2014). WB ; Mouse . PubMed: 24502509

Customer reviews and Q&As

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1-2 of 2 Abreviews or Q&A

Question

What are the species of the primary antibodies in this cocktail?

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Abcam community

Verified customer

Asked on Aug 01 2014

Answer

The primary antibodies in the primary cocktail were raised in either rabbit or mouse.

Read More

Kevin Hanson

Abcam Scientific Support

Answered on Aug 01 2014

Question

Inquiry:  For product ab140365, under "Applications"/"Membrane Fraction WB Cocktail images" the secondary antibody listed is "Secondary:HRP conjugated secondary antibody cocktail", is the secondary antibody cocktail included in ab140365?

Read More

Abcam community

Verified customer

Asked on Mar 04 2014

Answer

Yes, for ab140365 both the primary and secondary antibodies are included in the kit.

Read More

Abcam Scientific Support

Answered on Mar 04 2014

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