Overview

  • Product name
  • Description
    Rabbit polyclonal to S6K1
  • Host species
    Rabbit
  • Specificity
    Reacts strongly with 70 kd of p70 S6K, but only weakly with the 80 kDa S6K.
  • Tested applications
    Suitable for: WB, ELISA, IPmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Rabbit, Cow, Human, Pig
  • Immunogen

    Synthetic peptide corresponding to Human S6K1 (C terminal) conjugated to Keyhole Limpet Haemocyanin (KLH).
    Sequence:

    ISKRPEHLRMNL

  • Positive control
    • Use Mouse brain extract.

Properties

Applications

Our Abpromise guarantee covers the use of ab9366 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 1/250.
ELISA 1/500.
IP Use at 3 µg/mg of lysate.

Target

  • Function
    Acts to integrate nutrient and growth factor signals in regulation of protein synthesis, cell proliferation, cell growth, cell cycle progression and cell survival. Downstream effector of the mTOR signaling pathway. Phosphorylates specifically ribosomal protein S6 in response to insulin or several classes of mitogens. During translation initiation, the inactive form associatess with the eIF-3 complex under conditions of nutrient depletion. Mitogenic stimulation leads to phosphorylation and dissociation from the eIF-3 complex and the free activated form can phosphorylate other translational targets including EIF4B. Promotes protein synthesis by phosphorylating PDCD4 at 'Ser-67' and targeting it for degradation. Phosphorylates RICTOR leading to regulation of mammalian target of rapamycin complex 2 (mTORC2) signaling; probably phosphorylates RICTOR at 'Thr-1135'. Phosphorylates IRS1 at multiple serine residues coupled with insulin resistance; probably phosphorylates IRS1 at 'Ser-270'. Required for TNF-alpha induced IRS-1 degradation. Phosphorylates EEF2K in response to IGF1 and inhibits EEF2K activity. Phosphorylates BAD at 'Ser-99' in response to IGF1 leading to BAD inactivation and inhibition of BAD-induced apoptosis. Phosphorylates mitochondrial RMP leading to dissociation of a RMP:PPP1CC complex; probably phosphorylates RMP at 'Ser-99'. The free mitochondrial PPP1CC can dephosphorylate RPS6KB1 at Thr-412 which is proposed to be a negative feed back mechanism for the RPS6KB1 antiapoptotic function. Phosphorylates GSK3B at 'Ser-9' under conditions leading to loss of the TSC1-TSC2 complex. Phosphorylates POLDIP3.
  • Tissue specificity
    Widely expressed.
  • Sequence similarities
    Belongs to the protein kinase superfamily. AGC Ser/Thr protein kinase family. S6 kinase subfamily.
    Contains 1 AGC-kinase C-terminal domain.
    Contains 1 protein kinase domain.
  • Domain
    The autoinhibitory domain is believed to block phosphorylation within the AGC-kinase C-terminal domain and the activation loop.
    The TOS (TOR signaling) motif is essential for activation by mTORC1.
  • Post-translational
    modifications
    Phosphorylation at Thr-412 is regulated by mTORC1. The phosphorylation at this site is maintained by an agonist-dependent autophosphorylation mechanism.
  • Cellular localization
    Cytoplasm; Nucleus. Cytoplasm and Cell junction > synapse > synaptosome. Mitochondrion outer membrane.
  • Information by UniProt
  • Database links
  • Alternative names
    • 70 kDa ribosomal protein S6 kinase 1 antibody
    • KS6B1_HUMAN antibody
    • p70 alpha antibody
    • P70 beta 1 antibody
    • p70 ribosomal S6 kinase alpha antibody
    • p70 ribosomal S6 kinase beta 1 antibody
    • p70 S6 kinase alpha antibody
    • P70 S6 Kinase antibody
    • p70 S6 kinase, alpha 1 antibody
    • p70 S6 kinase, alpha 2 antibody
    • p70 S6K antibody
    • p70 S6K-alpha antibody
    • p70 S6KA antibody
    • p70(S6K) alpha antibody
    • p70(S6K)-alpha antibody
    • p70-alpha antibody
    • p70-S6K 1 antibody
    • p70-S6K antibody
    • P70S6K antibody
    • P70S6K1 antibody
    • p70S6Kb antibody
    • PS6K antibody
    • Ribosomal protein S6 kinase 70kDa polypeptide 1 antibody
    • Ribosomal protein S6 kinase beta 1 antibody
    • Ribosomal protein S6 kinase beta-1 antibody
    • Ribosomal protein S6 kinase I antibody
    • RPS6KB1 antibody
    • S6K antibody
    • S6K-beta-1 antibody
    • S6K1 antibody
    • Serine/threonine kinase 14 alpha antibody
    • Serine/threonine-protein kinase 14A antibody
    • STK14A antibody
    see all

References

This product has been referenced in:
  • Takeda K  et al. Activation of p70S6 Kinase-1 in Mesenchymal Stem Cells Is Essential to Lung Tissue Repair. Stem Cells Transl Med N/A:N/A (2018). Read more (PubMed: 29730892) »
  • Amara S  et al. Critical role of SIK3 in mediating high salt and IL-17 synergy leading to breast cancer cell proliferation. PLoS One 12:e0180097 (2017). Read more (PubMed: 28658303) »
See all 11 Publications for this product

Customer reviews and Q&As

1-10 of 16 Abreviews or Q&A

Answer

Thank you for contacting us.

Yes the customer would be eligible for a testing discount for ab12864 if they are willing to test it in mouse and/or sheep samples with western blotting. The sequence surrounding the Serine 235 and 236 of human RPS6 (SwissProt http://www.uniprot.org/uniprot/P62753), against which the antibody was raised, shares 100% homology with the mouse protein (SwissProt reference http://www.uniprot.org/uniprot/P62754) and we would therefore expect it to react. I was unable to find a sequence for the sheep protein but it seems that the sequence is highly conserved in mammals and I would therefore expect the antibody also to be able to detect the sheep protein.

In order for me to issue a testing discount could you please provide me with the email address and institute that the customer is with.

I look forward to receiving your reply.

Read More
Application
Western blot
Sample
Bos taurus Tissue lysate - whole (Liver)
Gel Running Conditions
Reduced Denaturing (4-15% Gradient)
Loading amount
20 µg
Specification
Liver
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Julie Kim

Verified customer

Submitted May 05 2017

Answer

Vielen Dank für Ihre Anfrage und dafür, dass Sie sich die Zeit genommen haben, unseren Fragebogen auszufüllen. Es tut mir leid, dass Sie Probleme mit unserem Antikörper hatten. Ich habe gesehen, dass Sie schon mit meiner Kollegin bezüglich des ab2571 in Kontakt waren. Haben Sie das Problem mit dem ab2571 gelöst? Wir würden uns sehr freuen zu hören, wie Sie vorangekommen sind.

In Bezug auf den ab9366,möchte ich gernedie folgenden Punkte ansprechen und Vorschlägezur Veränderungdes Protokolls machen.

1.) Können Sie bestätigen, dass der sekundäre Antikörper ansonsten gut funktioniert auf diesen Zelllysaten? Gibt der sekundäre Antikörper auch Hintergrundbanden?

2.) Ich kann Ihnen empfehlen, die Zelllysate in einem RIPA Puffer herzustellen. Die Wahl des Puffers ist wichtig, da einige Puffer gewisse Proteine nicht genug gut in Lösung bringen können. RIPA ist ein starker und guter Puffer. In unserem Onlineprotokoll haben wir auch einen Absatz über die Wahl der Puffer: https://www.abcam.com/index.html?pageconfig=resource&rid=11379#A1

3.) Ich kann auch empfehlen, Inhibitoren von Proteasen den Lysaten hinzufügen.

4.) Ich kann auch empfehlen, die Proben zu reduzieren. Falls wir keine spezielle Notiz auf dem Datenblatt haben, wurde der Antikörper unter reduzierten Kondition getestet.

Bitte lassen Sie mich wissen, ob ich Ihnen helfen konnte und zögern Sie nicht, sich wieder bei uns zu melden, falls Sie weitere Fragen haben.

Wie Sie wissen,garantieren wiralle Produkte so zu funktionieren wie auf dem Datenblatt beschrieben. Falls wir also das Problem nicht lösen können, werden wir Ihnen diesen Antikörperersetzen.

Ich freue mich wieder von Ihnen zu hören.

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Human Tissue lysate - whole (Liver)
Loading amount
30 µg
Specification
Liver
Gel Running Conditions
Reduced Denaturing (12)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C

Abcam user community

Verified customer

Submitted May 02 2012

Answer

Thank you for providing those details.


I can now confirm the discount code is xxxxx for which the expiration date is the 24th of July 2012. In order to activate the code, please submit an Abreview for ab12864 using mouse and/or sheep samples with western blotting, quoting the discount code in the "additional comments" section. More information on this offer can be found from the link:

https://www.abcam.com/collaboratordiscount

If you have any further questions, please do not hesitate to contact us again. I wish you all the best with your research.

Read More

Answer

Thank you for getting back to me.

I understand your concerns over the antibody. I suggested that the antibody may be at fault not because we have had other complaints from customers but because of the optimisation having been performed by the customer and the highly non-specific blot obtained. However, if you have used a further two antibodies to this target with similar results it may be that the samples do not contain the protein as expected. It may be if they have been stored at -80 for a long time, or gone through any freeze thaw cyclesthat they have degraded with time. It may be worthwhile to test the antibodies used with a positive control such as 293T cell lysate or jurkat cell lysate if you have these to hand? If you do not I would be able to provide this for you.

If you would prefer to take a replacement antibody, I can have this organised too. I have had a look at the antibodies suggested against RPS6. Both ab40820 and ab5961 seem like good choices against the unphosphorylated target. These antibodies have both been validated in western blotting and with mouse samples and would therefore both be covered by the Abpromise. From the antibodies chosen against the phosphorylated RPS6target, ab65748 and ab12864, I would suggest ab65748 would be the most appropriate to use as this has been validated with mouse samples which ab12864 has not (we can only say that we would expect it to work due to the sequence homology of the mouse protein with the immunogen used but we cannot guarantee it).

For all of the antibodies suggested, nonehave been used to detect the sheep protein and I cannot therefore say whether they would besuitable for performing western blottingwith thesesamples.

I hope this information has been of help. I look forward to hearing how you would like to proceed.

Read More

Answer

Thank you for taking time to complete our questionnaire and for providing me with that extra information. I am sorry 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. I appreciate the time you have spent in the laboratory and understand your concerns. It is regrettable the results have not been successful.

Having reviewed the protocol details, I believe this product should have given satisfactory results. It appears that you may have received a faulty vial. I apologize for the inconvenience has caused your customer and am pleased to offer you a free of charge replacement, credit note, or refund in compensation. Unfortunately, we only have the same lot of ab9366 in stock at the moment. I would therefore suggest https://www.abcam.com/S6K-antibody-6B2-ab119252.htmlwhich is a mouse monoclonal or https://www.abcam.com/S6K-antibody-ab47504.htmlwhich is a rabbit polyclonal. Both have been validated in WB and with mouse samples.

I would still be veryinterested in seeing the "no primary" control result, just to see if some improvements could be made in improving this non-specificity..

Thank you for your cooperation. I look forward to hearing from you with details of how you would like to proceed.

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Rat Cell lysate - whole cell (Liver)
Loading amount
50 µg
Specification
Liver
Gel Running Conditions
Reduced Denaturing (12%)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C

Abcam user community

Verified customer

Submitted Mar 13 2012

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Mouse Cell lysate - whole cell (Liver)
Loading amount
50 µg
Specification
Liver
Gel Running Conditions
Reduced Denaturing (12%)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C

Abcam user community

Verified customer

Submitted Mar 13 2012

Question

LOT NUMBER GR32217-3 ORDER NUMBER 978203 DESCRIPTION OF THE PROBLEM Wrong band size, possibly, multiple bands from 20-220KDa SAMPLE Mouse kidney tissue & lamb kidney tissue which had been snapo frozen at -80C PRIMARY ANTIBODY Ab9366. Rabbit polyclonal. 1/1000, 1/2000 diluted in PBST+2.5%FCS+0.01%azide. Incubated overnight at 4C. Wash in PBST 4 times for 15 minutes with shaking at room temp DETECTION METHOD ECL ANTIBODY STORAGE CONDITIONS 4C & -20C SAMPLE PREPARATION RIPA buffer + protease inhibitors, processed on ice/4C then frozen in aliquots at -80C AMOUNT OF PROTEIN LOADED 30ug ELECTROPHORESIS/GEL CONDITIONS 10% Reducing gel TRANSFER AND BLOCKING CONDITIONS Electro-transfer in TG + 0.1%SDS +20% methanol 2 hours at 4C. Blocking over 1hour in 5% low fat milk in PBST. SECONDARY ANTIBODY sigma goat anti-rabbit diluted 1/2000 up to 1/6000 in PBST. 2 hours room temp Wash in PBST 4 times for 15 minutes with shaking at room temp HOW MANY TIMES HAVE YOU TRIED THE APPLICATION? multi HAVE YOU RUN A "NO PRIMARY" CONTROL? Yes DO YOU OBTAIN THE SAME RESULTS EVERY TIME? Yes WHAT STEPS HAVE YOU ALTERED? Different blocking solutions (3%BSA instead of milk powder), block after primary, over night wash after secondary, numerous dilutions of primary & secondary. PVDF & nitrocellulose ADDITIONAL NOTES We have already spent some considerable time trying to resolve & optimize these antibodies as such we would appreciate your recommendation of antibodies that will work for our tissue. A publication is held up awaiting these result so there is urgency in this matter. As we have already used your antibodies without success we seek your advise before purchasing from another company. We have used S6K but if S6 would be a better option please let us know.

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. We take customer complaints very seriously and investigate every antibody we feel may not be performing as expected. 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 was hoping that you would be able to confirm the following details:

1. Could you share an image of the blot? This is hugely helpful in understanding the problems you have been experiencing.

2. You have mentioned that the antibody has been stored at both 4°C and -20°C. How was this performed, was it stored initially at 4°C then aliquotted to -20°C? From which storage conditions was the antibody used? Could you share images with me of the antibody used after storage at 4°C and when stored at 20°C?

3. You have mentioned that a "no-primary" control was performed. What was the results of this experiment? If possible, could you share an image of these results?

4. Could you share some more details of the secondary antibody used, the catalogue number would me most helpful.

5. Is the membrane being used fresh or has it been probed with a different antibodythen stripped? If so could you share the protocol used with me?

These details will allow me to more fully understand what may becontributing to the problems observed andfor me to make a more informed suggestion to you as to which antibody or protocol may work better for your samples.

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

1-10 of 16 Abreviews or Q&A

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