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  1. Link

    cell-fractionation-kit-standard-ab109719.pdf

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Cell Fractionation Kit - Standard (ab109719)

  • Datasheet
  • SDS
  • Protocol Booklet
Reviews (4)Q&A (37)References (82)

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Analysis of fractions by Western blot with a plasma membrane antibody cocktail.
  • Western blot of cytoplasmic, mitchondrial and nuclear fractions
  • Western blot

Key features and details

  • Assay type: Direct

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Overview

  • Product name

    Cell Fractionation Kit - Standard
    See all Cell Fractionation kits
  • Assay type

    Direct
  • Product overview

    Abcam's Cell Fractionation Kit (Standard) allows for the rapid and simple preparation of mitochondrial, cytoplasmic and nuclear containing fractions from cultured cells from mammals and other species.


    The Cell Fractionation Kit is designed to produce highly enriched fractions that allow for the monitoring of proteins of interest through various cellular compartments. It is not designed to produce purified cellular fractions.


    This kit does not require mechanical disruption of the sample. This is important as mechanical disruption can disrupt mitochondrial membranes, causing the biologically-irrelevant release of proteins. This kit is particularly useful in studying apoptosis and the movement of pro-apoptotic proteins such as cytochrome c.


    Sufficient materials are provided for fractionation of 1 x108 cells or for preparation of 40 samples, each corresponding to one 100 mm plate at 2.5 x 106 cells/plate.


    A companion product is also available which allows for the fractionation of cells grown in 96-well plates: Cell Fractionation Kit HT (ab109718).

  • Notes

     

Properties

  • Storage instructions

    Please refer to protocols.
  • Components 1 kit
    5X SDS Sample Buffer 1 x 10ml
    Buffer A 2x concentrate 1 x 175ml
    Detergent I 1 x 25µl
    Detergent II 1 x 1ml
  • Research areas

    • Kits/ Lysates/ Other
    • Kits
    • Molecular Biology Kits
    • Cell Fractionation Kits
    • Kits/ Lysates/ Other
    • Kits
    • Extraction Kit
    • Cell
  • Alternative names

    • MS862

Associated products

  • Related Products

    • Anti-Cytochrome C antibody [37BA11] (ab110325)
    • Cytochrome c Apoptosis ICC Antibody Kit (ab110417)

Images

  • Analysis of fractions by Western blot with a plasma membrane antibody cocktail.
    Analysis of fractions by Western blot with a plasma membrane antibody cocktail.

    Samples were loaded as follows from left to right: (1) Marker, (2) whole cell lysate, (3) cytosolic fraction lysate, (4) membrane fraction lysate and (5) nuclear fraction lysate. Membrane was blotted with a Plasma Membrane Fractionation WB cocktail ab140365 containing anti-Sodium Potassium ATPase antibody (110 kDa), anti-GAPDH antibody (37 kDa) and anti-Histone H3 (di methyl K9) antibody.

  • Western blot of cytoplasmic, mitchondrial and nuclear fractions
    Western blot of cytoplasmic, mitchondrial and nuclear fractions

    Cytosolic (C), mitochondrial (M) and nuclear (N) fractions of HepG2 cells were prepared as described in the Protocol. Fractions were analyzed by Western blotting using Abcam's ApoTrack™ Cytochrome c Apoptosis WB Antibody Cocktail (ab110415) containing antibodies against mitochondrial matrix (pyruvate dehydrogenase subunit E1α, PDH E1α), mitochondrial inner membrane (F1-ATPase α), mitochondrial intermembrane space (cytochrome c) and cytosolic (glyceraldehyde-3-phosphate dehydrogenase, GAPDH) markers as well as with antibodies against additional mitochondrial matrix (Hsp70) and nuclear (poly (ADP-ribose) polymerase, PARP and SP1) markers, followed by appropriate HRP-conjugated goat secondary antibodies and ECL detection.

  • Western blot
    Western blot

    In this experiment, apoptosis was induced in Jurkat, HeLa and 143B osteosarcoma cells by treatment with staurosporine or in Jurkat cells by FAS. Mitochondrial and cytoplasmic fractions were isolated (using ab109719) and probed using ab110415. As is clear from the gels, cytochrome c has translocated partially in FAS-induced cells and STS-treated osteosarcoma cells, and almost completely in STS-treated Jurkat and HeLa cells. The three control targets allow for verification of the "cleanness" of the cell fractionation.

Protocols

  • Protocol Booklet

Click here to view the general protocols

Datasheets and documents

  • SDS download

  • Datasheet download

    Download

References (82)

Publishing research using ab109719? Please let us know so that we can cite the reference in this datasheet.

ab109719 has been referenced in 82 publications.

  • Yeh CW  et al. The C-degron pathway eliminates mislocalized proteins and products of deubiquitinating enzymes. EMBO J 40:e105846 (2021). PubMed: 33469951
  • Aoto K  et al. ATP6V0A1 encoding the a1-subunit of the V0 domain of vacuolar H+-ATPases is essential for brain development in humans and mice. Nat Commun 12:2107 (2021). PubMed: 33833240
  • Fang X  et al. The HMGB1-AGER-STING1 pathway mediates the sterile inflammatory response to alkaliptosis. Biochem Biophys Res Commun 560:165-171 (2021). PubMed: 33992959
  • Duncan-Lewis C  et al. Cytoplasmic mRNA decay represses RNA polymerase II transcription during early apoptosis. Elife 10:N/A (2021). PubMed: 34085923
  • Lenis-Rojas OA  et al. Evaluation of the In Vitro and In Vivo Efficacy of Ruthenium Polypyridyl Compounds against Breast Cancer. Int J Mol Sci 22:N/A (2021). PubMed: 34445620
View all Publications for this product

Customer reviews and Q&As

Show All Reviews Q&A
Submit a review

Filter by Ratings

1-4 of 4 Abreviews

Cellular fractionation

Excellent Good 4/5 (Ease of Use)
Abreviews
Abreviews
abreview image
This kit was successfully used to separate the nuclear and cytoplasmic fractions. There was some contamination from the compartments so I would recommend an additional wash step, but overall the kit works well.

Abcam user community

Verified customer

Submitted Jul 14 2021

Excellent for nuclear extraction

Excellent Excellent 5/5 (Ease of Use)
Abreviews
Abreviews
abreview image
The kit was used to extract nuclear fraction from whole cell lysates. The mitochondrial fraction was not used. Lamin B1 was used as a nuclear marker and a strong signal was observed in the whole cell extract (WCE) and nuclear fractions (NUC) but not the cytoplasmic fraction (CYTO). Nuclear DNA was only sheared by vortexing and therefore the DNA present in the nuclear sample did cause smearing of the bands. Sonication could improve this but this was enough to demonstrate that the kit could quickly and efficiently purify nuclear fractions.

Abcam user community

Verified customer

Submitted Mar 31 2017

Got clean nuclear fraction

Good Excellent 5/5 (Ease of Use)
Abreviews
Abreviews
abreview image
The kit is quite easy to use. I got a clean nuclear fraction (N) with positive CREB staining and negative tubulin staining. However, the cytosolic fraction (C) has nuclear contamination (CREB positive).

DR. Simin Li

Verified customer

Submitted Dec 17 2016

Cell fractionation with Hela cells

Excellent Good 4/5 (Ease of Use)
Abreviews
Abreviews
abreview image
Follow the protocol in general, but wash with Buffer 1 between different extractions.

Abcam user community

Verified customer

Submitted Jan 14 2016

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