Anti-beta Actin antibody - Loading Control (ab8229)

Overview

  • Product nameAnti-beta Actin antibody - Loading ControlSee all beta Actin primary antibodies ...
  • Description
    Goat polyclonal to beta Actin - Loading Control
  • SpecificityThis antibody detects a single clean band at just above 40kD that corresponds to beta Actin.
  • Tested applicationsICC/IF, IHC-P, WBmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Rabbit, Chicken, Cow, Dog, Human, Fish, Monkey, Chinese Hamster
    Predicted to work with: a wide range of other species
  • Immunogen

    Synthetic peptide conjugated to KLH derived from within residues 1 - 100 of Human beta Actin.

    (Peptide available as ab13772.)

  • Positive control
    • In Western Blot, this antibody gave a positive signal in the following whole cell lysates: HeLa; NIH3T3; PC12; CHOK1. This antibody gave a positive result in IHC in the following FFPE tissue: Human normal colon.
  • General notes


    This antibody has been designed for use as a loading control and is ideal for this purpose.

Properties

Applications

Our Abpromise guarantee covers the use of ab8229 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
ICC/IF Use a concentration of 5 µg/ml.
IHC-P Use a concentration of 10 µg/ml. Perform heat mediated antigen retrieval via the pressure cooker method before commencing with IHC staining protocol.
WB 1/500. Detects a band of approximately 42 kDa (predicted molecular weight: 42 kDa).Can be blocked with Human beta Actin peptide (ab13772).

Target

Anti-beta Actin antibody - Loading Control images

  • All lanes : Anti-beta Actin antibody - Loading Control (ab8229) at 1/1000 dilution

    Lane 1 : HeLa whole cell (Human)
    Lane 2 : 3T3 cell (Mouse)
    Lane 3 : Rabbit Liver
    Lane 4 : MDCK cell (Dog)
    Lane 5 : EBTr cell (Cow)
    Lane 6 : SL-29 cell (Chicken)
    Lane 7 : CHO cell (Chinese Hamster)

    Lysates/proteins at 20 µg per lane.

    Secondary
    Rabbit Anti-Goat IgG H&L (HRP) (ab6741) at 1/5000 dilution
    developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 42 kDa
    Observed band size : 42 kDa


    Exposure time : 5 minutes
  • ICC/IF image of ab8229 stained human HeLa cells. The cells were methanol fixed (5 min), permabilised in TBS-T (20 min) and incubated with the antibody (ab8229, 5µg/ml) for 1h at room temperature. 1%BSA / 10% normal donkey serum / 0.3M glycine was used to quench autofluorescence and block non-specific protein-protein interactions. The secondary antibody (green) was Alexa Fluor® 488 donkey anti-goat IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red). DAPI was used to stain the cell nuclei (blue).

  • IHC image of beta Actin staining in Human normal colon formalin fixed paraffin embedded tissue section*. The section was pre-treated using pressure cooker heat mediated antigen retrieval with sodium citrate buffer (pH6) for 30mins. The section was incubated with ab8229, 10µg/ml overnight at +4°C. An HRP-conjugated secondary (Ab6885, Donkey polyclonal to Goat IgG - H&L (HRP) 1/250 dilution) was used for 1hr at room temperature. The section was counterstained with haematoxylin and mounted with DPX.

    *Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

  • Lanes 1 - 4 : Anti-beta Actin antibody - Loading Control (ab8229) at 1 µg/ml (Blocked with 5% BSA.)
    Lanes 5 - 8 : Anti-beta Actin antibody - Loading Control (ab8229) at 1 µg/ml (Blocked with 3% Milk.)

    Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
    Lane 2 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate
    Lane 3 : PC12 (Rat adrenal pheochromocytoma cell line) Whole Cell Lysate
    Lane 4 : CHO-K1 cell lysate Whole Cell Lysate
    Lane 5 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
    Lane 6 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate
    Lane 7 : PC12 (Rat adrenal pheochromocytoma cell line) Whole Cell Lysate
    Lane 8 : CHO-K1 cell lysate Whole Cell Lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    Rabbit polyclonal to Goat IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution
    developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 42 kDa
    Observed band size : 42 kDa
    Additional bands at : 48 kDa. We are unsure as to the identity of these extra bands.

    Exposure time : 20 minutes

    This blot was produced using a 10% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using 5% Bovine Serum Albumin (lanes 1-4) or 3% milk (lanes 5-8) before being incubated with ab8229 overnight at 4°C. Antibody binding was detected using an anti-Goat antibody conjugated to HRP, and visualised using ECL development solution.  A non-specific band at 50-kDa is observed on western blots of some of our batches of ab8229. We have found that milk blocking is more effective than BSA blocking in eliminating this non-specific binding. 

References for Anti-beta Actin antibody - Loading Control (ab8229)

This product has been referenced in:
  • Hall J  et al. Evolution of a developmental mechanism: Species-specific regulation of the cell cycle and the timing of events during craniofacial osteogenesis. Dev Biol 385:380-95 (2014). WB . Read more (PubMed: 24262986) »
  • Röhrl C  et al. Endoplasmic reticulum stress impairs cholesterol efflux and synthesis in hepatic cells. J Lipid Res 55:94-103 (2014). WB . Read more (PubMed: 24179149) »

See all 32 Publications for this product

Product Wall

Application Western blot
Loading amount 25 µg
Gel Running Conditions Reduced Denaturing (10)
Sample Human Cell lysate - whole cell (HEK293)
Specification HEK293
Blocking step (agent) for 1 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: 20°C
Username

Abcam user community

Verified customer

Submitted May 07 2014

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Mouse Cell lysate - whole cell (neuronal)
Loading amount 30 µg
Specification neuronal
Gel Running Conditions Reduced Denaturing (10)
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2%
Username

Dr. Nicola Bright

Verified customer

Submitted Nov 28 2012

Thank you for confirming these details and for your cooperation. The details provided enable us to closely monitor the quality of our products.

I am sorry this product did not perform as stated on the datasheet and for the inconvenience this h...

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Thank you for your email. I am sorry to hear that you have been experiencing problems with this antibody.

I have read the details you have kindly provided and have following further questions for better understanding of the problem;

-...

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Thank you for contacting us.

Here are the links for the MSDS for the 2 products:

For ab6741: http://www.abcam.com/ps/Products/6/ab6741/Msds/ab6741_WPNA_EN.pdf
For ab8229: http://www.abcam.com/ps/Products/8/ab8229/Msds/ab8229_WPNA_...

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Thank you for your reply.

We do not currently have any other lots of ab8229 available. We have primary antibodies against actin that were raised in mouse and chicken. Would one of those work better for you?

Please let me know how yo...

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Thank you for your reply.

I was wondering if ab8227 would be a good replacement for you:

http://www.abcam.com/beta-Actin-antibody-Loading-Control-ab8227.html



If it is, then please let me know and I will get it s...

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Thank you for contacting us.

I am sorry that this antibody did not perform as stated on the datasheet, As requested, I have asked our Finance department to issue a credit note for you.

Credit ID: 20043

The credit note m...

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Thank you for your message. I am sorry to hear the free of charge replacement is also not providing satisfactory results and that this antibody is not working for you.

I appreciate the time you have spent on these experiments.I would suggest...

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Thank you for confirming these details and for your cooperation. The details provided enable us to closely monitor the quality of our products.

I am sorry this product did not perform as stated on the datasheet and for the inconvenience this...

Read More

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"