Overview

  • Product name
  • Description
    Rabbit polyclonal to pan-AKT
  • Host species
    Rabbit
  • Tested applications
    Suitable for: ICC, IHC-P, WB, ICC/IF, IHC-Fr, Flow Cyt, ELISAmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
    Predicted to work with: Chicken
  • Immunogen

    Synthetic peptide:

    CRPHFPQFSYSASGTA

    conjugated to KLH, corresponding to amino acids 466-480 of Human pan-AKT (100% similar to Rat, Chicken and Mouse AKT sequences). (Peptide available as ab9041.)

  • Positive control
    • WB: NIH/3T3 whole cell lysate IHC: colon tumour sections

Properties

Applications

Our Abpromise guarantee covers the use of ab8805 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 1/80.
IHC-P 1/1000 - 1/1500.
WB 1/500. Detects a band of approximately 60 kDa (predicted molecular weight: 56 kDa).
ICC/IF Use at an assay dependent concentration.
IHC-Fr Use at an assay dependent concentration. PubMed: 24759991
Flow Cyt Use at an assay dependent concentration.
ELISA 1/2000 - 1/10000.

Target

  • Function
    Plays a role as a key modulator of the AKT-mTOR signaling pathway controlling the tempo of the process of newborn neurons integration during adult neurogenesis, including correct neuron positioning, dendritic development and synapse formation (By similarity). General protein kinase capable of phosphorylating several known proteins. Phosphorylates TBC1D4. Signals downstream of phosphatidylinositol 3-kinase (PI(3)K) to mediate the effects of various growth factors such as platelet-derived growth factor (PDGF), epidermal growth factor (EGF), insulin and insulin-like growth factor I (IGF-I). Plays a role in glucose transport by mediating insulin-induced translocation of the GLUT4 glucose transporter to the cell surface. Mediates the antiapoptotic effects of IGF-I. Mediates insulin-stimulated protein synthesis by phosphorylating TSC2 at 'Ser-939' and 'Thr-1462', thereby activating mTORC1 signaling and leading to both phosphorylation of 4E-BP1 and in activation of RPS6KB1. Promotes glycogen synthesis by mediating the insulin-induced activation of glycogen synthase. The activated form can suppress FoxO gene transcription and promote cell cycle progression. Essential for the SPATA13-mediated regulation of cell migration and adhesion assembly and disassembly.
  • Tissue specificity
    Expressed in all human cell types so far analyzed. The Tyr-176 phosphorylated form shows a significant increase in expression in breast cancers during the progressive stages i.e. normal to hyperplasia (ADH), ductal carcinoma in situ (DCIS), invasive ductal carcinoma (IDC) and lymph node metastatic (LNMM) stages.
  • Involvement in disease
    Defects in AKT1 are a cause of susceptibility to breast cancer (BC) [MIM:114480]. A common malignancy originating from breast epithelial tissue. Breast neoplasms can be distinguished by their histologic pattern. Invasive ductal carcinoma is by far the most common type. Breast cancer is etiologically and genetically heterogeneous. Important genetic factors have been indicated by familial occurrence and bilateral involvement. Mutations at more than one locus can be involved in different families or even in the same case.
    Defects in AKT1 are associated with colorectal cancer (CRC) [MIM:114500].
    Defects in AKT1 are associated with susceptibility to ovarian cancer [MIM:604370]; also called susceptibility to familial breast-ovarian cancer type 1 (BROVCA1).
  • Sequence similarities
    Belongs to the protein kinase superfamily. AGC Ser/Thr protein kinase family. RAC subfamily.
    Contains 1 AGC-kinase C-terminal domain.
    Contains 1 PH domain.
    Contains 1 protein kinase domain.
  • Domain
    Binding of the PH domain to the phosphatidylinositol 3-kinase alpha (PI(3)K) results in its targeting to the plasma membrane. The PH domain mediates interaction with TNK2 and Tyr-176 is also essential for this interaction.
    The AGC-kinase C-terminal mediates interaction with THEM4.
  • Post-translational
    modifications
    Phosphorylation on Thr-308, Ser-473 and Tyr-474 is required for full activity. Activated TNK2 phosphorylates it on Tyr-176 resulting in its binding to the anionic plasma membrane phospholipid PA. This phosphorylated form localizes to the cell membrane, where it is targeted by PDPK1 and PDPK2 for further phosphorylations on Thr-308 and Ser-473 leading to its activation. Ser-473 phosphorylation by mTORC2 favors Thr-308 phosphorylation by PDPK1. Ser-473 phosphorylation is enhanced by interaction with AGAP2 isoform 2 (PIKE-A). Ser-473 phosphorylation is enhanced in focal cortical dysplasias with Taylor-type balloon cells.
    Ubiquitinated; undergoes both 'Lys-48'- and 'Lys-63'-linked polyubiquitination. TRAF6-induced 'Lys-63'-linked AKT1 ubiquitination is critical for phosphorylation and activation. When ubiquitinated, it translocates to the plasma membrane, where it becomes phosphorylated. When fully phosphorylated and translocated into the nucleus, undergoes 'Lys-48'-polyubiquitination catalyzed by TTC3, leading to its degradation by the proteasome.
  • Cellular localization
    Cytoplasm. Nucleus. Cell membrane. Nucleus after activation by integrin-linked protein kinase 1 (ILK1). Nuclear translocation is enhanced by interaction with TCL1A. Phosphorylation on Tyr-176 by TNK2 results in its localization to the cell membrane where it is targeted for further phosphorylations on Thr-308 and Ser-473 leading to its activation and the activated form translocates to the nucleus.
  • Information by UniProt
  • Database links
  • Alternative names
    • AKT1 antibody
    • AKT1_HUMAN antibody
    • AKT2 antibody
    • AKT3 antibody
    • PKB alpha antibody
    • PKB antibody
    • PKB beta antibody
    • PKBalpha antibody
    • PRKBA antibody
    • PRKBB antibody
    • PRKBG antibody
    • Protein kinase B antibody
    • Protein kinase B beta antibody
    • Protein kinase B gamma antibody
    • Proto-oncogene c-Akt antibody
    • RAC alpha antibody
    • RAC antibody
    • RAC gamma antibody
    • RAC PK alpha antibody
    • RAC PK beta antibody
    • RAC PK gamma antibody
    • RAC-alpha serine/threonine-protein kinase antibody
    • RAC-PK-alpha antibody
    • STK2 antibody
    see all

Images

  • Western blot using Akt antibody ab8805 at 1/500, Goat anti-rabbit IgG peroxidase conjugate at 1/10,000. 20 ug NIH/3T3 whole cell lysate, Gel type 10% NuPage with MOPS buffer developed with Substrate Pierce SuperSignal™ West Pico

    Western blot using Akt antibody ab8805 at 1/500, Goat anti-rabbit IgG peroxidase conjugate at 1/10,000. 20 ug NIH/3T3 whole cell lysate, Gel type 10% NuPage with MOPS buffer developed with Substrate Pierce SuperSignal™ West Pico
  • The Akt antibody (ab8805) is used at 1/80 on cultured neonatal rat cardiomyocytes that express a nuclear-targeted Akt construct. The green is Akt antibody, the red is Texas-red™ phalloidin that labels actin filaments.

  • A: Normal colon tissue
    B: Tumour tissue
    Akt antibody (ab8805) used at 1/1000 on formalin-fixed paraffin embedded tissue.

  • Anti-pan-AKT antibody (ab8805) at 1/500 dilution + Lysate prepared from human HT1080 cell line at 10 µg

    Secondary
    HRP-conjugated donkey polyclonal to rabbit IgG at 1/5000 dilution

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 56 kDa
    Observed band size: 57 kDa
    why is the actual band size different from the predicted?


    Exposure time: 6 minutes

    See Abreview

Protocols

References

This product has been referenced in:
  • Pi Z  et al. Isoflurane reduces pain and inhibits apoptosis of myocardial cells through the phosphoinositide 3-kinase/protein kinase B signaling pathway in mice during cardiac surgery. Mol Med Rep 17:6497-6505 (2018). Read more (PubMed: 29488606) »
  • Wang L  et al. TIPE-2 suppresses growth and aggressiveness of hepatocellular carcinoma cells through downregulation of the phosphoinositide 3-kinase/AKT signaling pathway. Mol Med Rep 17:7017-7026 (2018). Read more (PubMed: 29568863) »
See all 121 Publications for this product

Customer reviews and Q&As

1-10 of 24 Abreviews or Q&A

Abcam has not validated the combination of species/application used in this Abreview.
Application
Western blot
Sample
Bos taurus Cell lysate - other (HEK293T + BoMAC (bovine macrophage))
Gel Running Conditions
Reduced Denaturing (10% SDSPAGE)
Loading amount
15 µg
Treatment
1% triton-X-100. P=insoluble fraction, S=soluble lysate
Specification
HEK293T + BoMAC (bovine macrophage)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 18°C

Abcam user community

Verified customer

Submitted Mar 08 2016

Abreviews
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Loading amount
20 µg
Gel Running Conditions
Reduced Denaturing (4-15 % NuPAGE)
Sample
Human Cell lysate - whole cell (Endomteriotic)
Specification
Endomteriotic
Blocking step
BSA as blocking agent for 16 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 4°C

Dr. Brett Mckinnon

Verified customer

Submitted Jun 10 2013

Answer

We do!

For the pan-Akt antibody we have ab4702, colon tumor slides, and ab139513 which is NIH3T3 cell lysate.

The for the Akt1 pS473 antibody ab66138 we have human breast cancer tissue slides ab46961, and TNF treated NIH3T3 lysate ab14877.

We are continually trying to add controls to our catalog but sometimes they are harder to find so please feel free to contact us should you ever have difficulty finding a product!

Read More

Answer

Thank you for taking time to complete our questionnaire. I am sorry to hear that this antibody is not providing satisfactory results.

The details provided will enable us to investigate this case and will provide us with vital information for monitoring product quality.

Having reviewed this case, I would like to offer some suggestions to help optimize the results for these product. I would also appreciate if you can confirm some further details:

As each lane was running the same sample we should expect to see a uniform pattern for each. However in the image provided this is definitly not true. This issue is most commonly found with a bad gel. Sometimes they may go bad if stored incorrectly or for too long but most often the cause is that is is run at too high a voltage or with running buffers that have gone bad. It also seems that an 8% gel may be a less than ideal choice for proteins of this size. I recommend 12% SDS-PAGE for these.

I would suggest using new or commercial gels of 12%, running with fresh running buffers at a lower voltage. I would also suggest using BSA 3% in PBS + 0.1% Tween-20 as a blocking buffer for ab8805 and ab66127 and prehaps a casein based blocking buffer such as ab64224https://www.abcam.com/Protein-Block-ab64226.htmlfor use with the phospho-S473 AKT1 antibody (ab66138).

Should the suggestions not improve the results, please do let me know.

In the event that a product is not functioning in the species and applications cited on the product datasheet (and the problem has been reported within 6 months of purchase), we would be pleased to provide a free of charge replacement, credit note, or refund.

I hope this information is helpful, and I thank you for your cooperation.

Read More

Answer

Thank you for contacting us.

I am sorry to hear you have been experiencing problems with one of our antibodies. The quality of our products is important to us and I would like to reassure you that we investigate all customer complaints. I have had the opportunity to review the information and images that you have provided and would like to ask a few questions and offer some suggestions that may help.

These products are covered by our Abpromise guarantee.If we cannot remedy this issue and this is a product that you have purchased within the last six months, we will replace or refund it under our Abpromise guarantee, as you are using it according to specifications listed on our datasheet.

In regards each image, were the samples in each lane the same? Were they different animals, protein concentration? These blots in images for ab8805 and ab66138 appear to have not run through and the gel in the image for ab66127 look like it may have deformed during running. Do you happen to have whole gel images for any of these?

The quality of western blots using PVDF membranes will be greatly increased by pre-wetting the membrane in methanol.If this is not something that you have already dome I would suggest doing this. PVDF is hydrophobic and therefore should be prewet in methanol before it is used. Wet the membrane in methanol for 15 seconds. Membrane should uniformly change from opaque to semi-transparent. Carefully place the membrane in ultrapure water and soak for 2 minutes. Then carefully place the membrane in transfer buffer and let equilibrate for at least 5 minutes. You should get a cleaner signal with less antibody when performing this.

I hope that this information is helpful. Please let me know if you have any questions or there are other ways that Abcam may help you meet your research goals.

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Mouse Cell lysate - whole cell (macrophage cell lysate)
Loading amount
20 µg
Specification
macrophage cell lysate
Gel Running Conditions
Reduced Denaturing
Blocking step
Milk as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Mar 09 2012

Answer

Thank you for contacting us. I would say that this sort of experiment is very-very tricky. We unfortunately do not have any protocol available at the moment for manually IHC staining quantification with antibodies. I would suggest ploughing through the publications and finding if there are scientists who are using similar kind of method. http://www.histochem.org/archives/photoshop.pdf http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1810239/ http://www.ncbi.nlm.nih.gov/pubmed/21370029 Have a look at these articles and check which one could help you to set up the experiment. I hope this will help.

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Sample
Mouse Cell (Mouse embryonic stem cells)
Specification
Mouse embryonic stem cells
Fixative
4% PFA
Permeabilization
Yes - 0.1% Triton
Blocking step
Serum as blocking agent for 30 minute(s) · Concentration: 10% · Temperature: RT°C

Abcam user community

Verified customer

Submitted Nov 23 2010

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Mouse Cell lysate - whole cell (NIH 3T3 cell line)
Loading amount
10 µg
Specification
NIH 3T3 cell line
Gel Running Conditions
Reduced Denaturing (10%)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C

Abcam user community

Verified customer

Submitted Nov 13 2009

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Human Cell lysate - whole cell (HT1080 cell line)
Loading amount
10 µg
Specification
HT1080 cell line
Gel Running Conditions
Reduced Denaturing (10%)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C

Abcam user community

Verified customer

Submitted Nov 13 2009

1-10 of 24 Abreviews or Q&A

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