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Mitochondria Fraction Western Blot Cocktail (ab139416)

Key features and details

  • Sample type: Adherent cells, Cell culture extracts, Cell Lysate, Suspension cells, Tissue Extracts, Tissue Homogenate

Overview

  • Product name

    Mitochondria Fraction Western Blot Cocktail

  • Sample type

    Cell culture extracts, Adherent cells, Suspension cells, Tissue Extracts, Cell Lysate, Tissue Homogenate

  • Species reactivity

    Reacts with: Mouse, Rat, Human

  • Product overview

    Mitochondria Fraction Western Blot Cocktail (ab139416) contains 3 mAbs each targeting a specific organelle marker. The presence of mitochondria is determined by Anti-ATP5A ; cytosol by Anti-GAPDH; and nucleus by Anti-Histone H3 (di methyl K9). This cocktail is suitable for determining the purity of organelle isolates prior to further characterization.


     


    This product is particularly valuable to researchers working in organelle proteomics. Mass spectrometry is frequently used in this field to determine the protein content of targeted organelle isolates. These isolates are obtained using differential centrifugation, density gradient fractionation, biochemical enrichment, or affinity purification. Unfortunately, the various methods of purification available for organelle isolation are imperfect and leave behind contaminants from undesired regions of the cell. These contaminants are inevitable, but being aware of which contaminants are present is crucial for analysis of mass spectrometry results. The high sensitivity and species cross reactivity of the antibodies in this cocktail will quickly and easily reveal impurities caused by imperfect sample preparation.

  • Tested applications

    Suitable for: WBmore details

Properties

Images

  • Western Blot for Mitochondrial Antibody Cocktail – Component Separation
    Western Blot for Mitochondrial Antibody Cocktail – Component Separation
    Developed using the ECL technique; Performed under reducing conditions; Exposure time: 5 mins ; All blocking and antibody incubation steps were done in 5% milk, 20 mM Tris-HCl, 0.1% TWEEN-20

    Sample 1: Marker
    Samples 2-5: MHH (Mouse heart homogenate) Whole Tissue Lysate – 20 µg

    Primary:
    Lane 1: none
    Lane 2: Anti- ATP5A antibody – Mitochondrial Membrane Marker
    Lane 3: Anti- GAPDH antibody – Cytosolic Membrane Marker
    Lane 4: Anti-Histone H3 (di methyl K9) antibody – Nuclear Membrane Marker
    Lane 5: Assembled Mitochondrial Membrane Antibody Cocktail

    Secondary:ab131368 at 1/1000 dilution

    Predicted ATP5a band size: 60 kDa
    Observed ATP5a band size: 55 kDa

    Predicted GAPDH band size: 37 kDa
    Observed GAPDH band size: 37 kDa

    Predicted Histone H3 (di methyl K9) band size: 17 kDa
    Observed Histone H3 (di methyl K9) band size: 17 kDa
  • Western Blot for Mitochondrial Antibody Cocktail – HeLa Fractions
    Western Blot for Mitochondrial Antibody Cocktail – HeLa Fractions
    Sample Preparation: HeLa cell lysate prepared using the Membrane Fractionation Kit (ab139409); Developed using the ECL technique under reducing conditions; Exposure time: 5 mins; Blocking and antibody incubation steps done in 5% milk, 20 mM Tris-HCl, 0.1% TWEEN-20

    Lane 1: Marker
    Lane 2 : HeLa Whole Cell Lysate - 20 µL
    Lane 3 : HeLa Cytosolic Fraction Lysate - 20 µL
    Lane 4 : HeLa Membrane Fraction Lysate - 20 µL
    Lane 5 : HeLa Nuclear Fraction Lysate - 20 µL

    All Lanes:
    Anti- ATP5A antibody – Mitochondrial Marker – 1/250 dilution
    Anti- GAPDH antibody – Cytosolic Marker – 1/250 dilution
    Anti-Histone H3 (di methyl K9) antibody – Nuclear Marker – 1/250 dilution

    Secondary: Goat polyclonal to Mouse IgG (ab6789) – H&L – Pre-Absorbed (HRP) at 1/10000 dilution

    Observed ATP5a band size: 55 kDa
    Observed GAPDH band size: 37 kDa
    Observed Histone H3 (di methyl K9) band size: 17 kDa
  • Western Blot for Mitochondrial Membrane Antibody Cocktail – Cross Reactivity
    Western Blot for Mitochondrial Membrane Antibody Cocktail – Cross Reactivity
    Developed using the ECL technique; Performed under reducing conditions; Exposure time: 3 mins; All blocking and antibody incubation steps done in 5% milk, 20 mM Tris-HCl, 0.1% TWEEN-20

    Sample 1: Marker
    Sample 2: HHH Whole Tissue Lysate - 20 µg
    Sample 3: Hela Whole Cell Lysate – 20 µg
    Sample 4: MHH Whole Tissue Lysate - 20 µg
    Sample 5: NIH3T3 Whole Cell Lysate – 20 µg
    Sample 6: RHH Whole Tissue Lysate – 20 µg
    Sample 7: H9C2 Whole Cell Lysate – 20 µg

    All Lanes:
    Anti- ATP5A antibody – Mitochondrial Marker – 1/250 dilution
    Anti- GAPDH antibody – Cytosolic Marker – 1/250 dilution
    Anti-Histone H3 (di methyl K9) antibody – Nuclear Marker – 1/250 dilution

    Secondary: ab131368 at 1/1000 dilution

    Observed ATP5A band size: 60 kDa
    Observed GAPDH band size: 37 kDa
    Observed Histone H3 (di methyl K9) band size: 15 kDa

Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
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