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  1. Link

    products/cell-lysates/nih3t3-whole-cell-lysate-ab7179.pdf

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NIH/3T3 whole cell lysate (ab7179)

  • Datasheet
  • SDS
Submit a review Q&A (5)

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Overview

  • Product name

    NIH/3T3 whole cell lysate
    See all NIH 3T3 lysates
  • General notes

    Cell line: NIH/3T3 (Mouse embryonic fibroblast).
    Growth media: DMEM and 10% bovine calf serum.


    Mouse NIH/3T3 cell lysate was prepared by homogenization in modified RIPA buffer(50 mM Tris-HCl, pH 7.4, 1% Triton X-100, 0.2% sodium deoxycholate, 0.2% sodium dodecylsulfate (SDS), 1 mM sodium ethylenediaminetetraacetate,1 mM phenylmethyl-sulfonyl flouride, 5 µg/ml of aprotinin, 5 µg/ml of leupeptin). Cell debris was removed by centrifugation. Protein concentration was determined with Bio-Rad protein assay. The lysate was boiled for 5 min in 1 x SDS sample buffer (50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1% SDS, 0.01% bromophenol blue) containing 5% b-mercaptoethanol.

    Abcam has not and does not intend to apply for the REACH Authorisation of customers’ uses of products that contain European Authorisation list (Annex XIV) substances.
    It is the responsibility of our customers to check the necessity of application of REACH Authorisation, and any other relevant authorisations, for their intended uses.

  • Tested applications

    Suitable for: WBmore details

Properties

  • Mycoplasma free

    Yes
  • Form

    Liquid
  • Storage instructions

    Shipped at 4°C. Upon delivery aliquot. Store at -80°C. Avoid freeze / thaw cycle.
  • Storage buffer

    pH: 7.2
    Constituents: 60.05% Water, 12.5% Glycerol (glycerin, glycerine), 9% Tris HCl, 7.7% DTT, 4.4% Sodium chloride, 1% Triton-X-100, 1% Sodium deoxycholate, 1.1% Sodium lauryl sulfate, 0.15% EDTA disodium salt, 0.5% Aprotinin, 0.5% Leupeptin hemisulfate, 0.09% PMSF, 0.01% Bromophenol blue
  • Concentration information loading...
  • Lysate notes

    Mouse NIH/3T3 cell lysate was prepared by homogenization in modified RIPA buffer(50 mM Tris-HCl, pH 7.4, 1% Triton X-100, 0.2% sodium deoxycholate, 0.2% sodium dodecylsulfate (SDS), 1 mM sodium ethylenediaminetetraacetate,1 mM phenylmethyl-sulfonyl flouride, 5 µg/ml of aprotinin, 5 µg/ml of leupeptin). Cell debris was removed by centrifugation. Protein concentration was determined with Bio-Rad protein assay. The lysate was boiled for 5 min in 1 x SDS sample buffer (50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1% SDS, 0.01% bromophenol blue) containing 5% b-mercaptoethanol.
  • Research areas

    • Kits/ Lysates/ Other
    • Lysates
    • Whole Cell Lysates
    • Mouse
    • Other
  • Background

    NIH 3T3 cells are established from a NIH Swiss mouse embryo. These cells are highly contact inhibited and are sensitive to sarcoma virus focus formation and leukaemia virus propagation. Cells have now lost their contact inhibition. This cell line was established from NIH Swiss mouse embryo cultures in the same manner as the original random bred 3T3 and the inbred BALB/c 3T3. The established NIH/3T3 line was subjected to more than 5 serial cycles of subcloning in order to develop a subclone with morphologic characteristics best suited for transformation assays. It is therefore used for DNA transfection studies

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab7179 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
Use at an assay dependent dilution.
NIH/3T3 cell lysate is ready to load on SDS-PAGE for Western blotting, 20 µg per lane is recommended for mini gel.
Notes
WB
Use at an assay dependent dilution.

NIH/3T3 cell lysate is ready to load on SDS-PAGE for Western blotting, 20 µg per lane is recommended for mini gel.

Protocols

To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.

Click here to view the general protocols

Datasheets and documents

  • SDS download

  • Datasheet download

    Download

References (0)

Publishing research using ab7179? Please let us know so that we can cite the reference in this datasheet.

ab7179 has not yet been referenced specifically in any publications.

Customer reviews and Q&As

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1-5 of 5 Abreviews or Q&A

Question

On the basis of the result of microarray , it is reasonable that the upper band means p19 expression.

But the positive control (NIH3T3) located on the lower band. Moreover , the band size seems to be a little bit lower than 19 kDa.

I don’t know which band shows the actual p19. Also, is there any possibility that the location of the band is different between samples and NIH3T3 ?

Read More

Abcam community

Verified customer

Asked on Oct 30 2012

Answer

Thank you for contacting us.

Based on this image alone it seems like the lower band is your CDKN2A/p19ARF band. It is possible for band size to change slightly due to sample preparation or buffer, however usually the positive control will be a good indicator of where the protein of interest is. One strategy to confirm the band might be to use the blocking peptide (the peptide used as the immunogen) to specifically block the CDKN2A/p19ARF protein. We have this available as ab
2659.


I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

Use our products? Submit an Abreview. Earn rewards!
https://www.abcam.com/abreviews

Read More

Abcam Scientific Support

Answered on Oct 30 2012

Question

Hello,

Yesterday, I used your p19ARF antibody(ab80) for my samples. Also I set NIH3T3 whole cell lysate as a positive control, but it didn’t work. (no band.)

I borrowed this cell line from other lab. After I cultured, I extracted lysate by myself.

I think there is some possibility that my NIH3T3 had any problems. So, I plan to order your cell lysate and check some mouse embryonic fibroblast cell line; ab7179,ab14874,ab7901,ab3874.

I’d like to ask you which is the best as positive control for p19ARF antibody? Or is there no difference among them?

Thank you,

Read More

Abcam community

Verified customer

Asked on Oct 11 2012

Answer

Thank you very much for contacting us with your enquiry.

I'm sorry to hear that there were no bands in Western blot using this antibody. I would expect any of the MEF cell lines to work as a positive control, but I suggest using a whole cell lysate (ab14874 is a nuclear fraction) since that is what we have tested with ab80. What kind of samples are you using other than the MEF lysate?

If you'd like to send some more details about your protocol ( how much protein was loaded, dilution of antibody, blocking solution, etc) I'd be happy to take a look and see if we have any suggestions to improve the results.

I look forward to hearing from you. If there's anything else that we can do for you, please let me know and I'll be happy to help.

Read More

Abcam Scientific Support

Answered on Oct 11 2012

Question

I'm having trouble using all abcam products with alpha smooth muscle actin antibody, ab 5694. I have attached a picture showing a ladder, skeletal muscle whole cell lysate (ab3869), and NIH3T3 whole cell lysate (ab7179). I loaded 10ug of protein into each well, 5% BSA block overnight, 2 hour of primary at 1ug/mL in 10mL TBS-T, 1:15000 secondary goat anti rabbit (ab97051), visualized with ECL+. 2 questions: 1) What muscle is the skeletal muscle lysate coming from, and should it have alpha-SMA? and 2) Why is my NIH3T3 lane not staining for alpha-SMA? Thanks!

Read More

Abcam community

Verified customer

Asked on Jul 06 2012

Answer

I apologize for the delay in getting back to you. I am still waiting for a reply from our source of the skeletal muscle lysate. I do not think it should contain alpha-SMA.

On the other hand, the fault may be with the antibody, if is not detecting this protein in the NIH3T3 lysate. I suspect that the antibody, though we have not received other complaints about it recently, is cross-reacting with something in the skeletal lysate that happens to be the same size as SMA.

Have you tried staining blots of these lysates with any other antibodies? If so, were the results as expected?

Read More

Abcam Scientific Support

Answered on Jul 06 2012

Question

I am interested in purchasing a fibroblast whole cell lysate product from you company to test a new procollagen probe my lab developed.

I noticed that you sell several kinds of fibroblast lysate products:ab7179, ab3960, ab3874, ab7901...

I wanna know if these products have a high procollagen (˜150KDa) content. Have you tested that? Or do you know if there have been any existing reports in which these products have been used for procollagen detection? This information is important for me to make the decision.

Read More

Abcam community

Verified customer

Asked on Apr 10 2012

Answer

Thank you for contacting Abcam and for your interest in our whole cell lysates.

Regarding these products, they are not tested for protein composition. They are only tested for protein concentration and confirmed to not be degraded.

If you are interested in determining which lysates will contain the protein that you need, I would recommend a search of the literature. Unfortunately, we are not experts in this field and would not want to recommend a product that is not suitable for your experiments.

I hope this information is helpful. Please do not hesitate to contact us if you have any additional questions.

Read More

Abcam Scientific Support

Answered on Apr 10 2012

Question

I just recieved my shipment. The label says the vial holds 100ug at (2mg/mL). The data sheet suggests that I load 20 ug per lane as a control for POMC. The vial actually contained 100 uL. So, is the concentration correct and the 100ug is incorrect or vice versa? I need to know what volume of this lysate would be equal to 20ug of protein.

Read More

Abcam community

Verified customer

Asked on Aug 29 2006

Answer

Thank you for your enquiry. You are correct, this antibody is at a concentration of 2mgml-1. The antibody is provided at 100 µg therefore 50ul. We do receive some very occasional shortages which requires our lab team taking a vial and adding lysate to the other vials that are short of antibody. This can result in customers receiving more antibody or lysate than expected. I would recommend that you prepare 20ug (20ul) of lysate. I hope this information helps, please do not hesitate to contact us if you need any more advice or information.

Read More

Abcam Scientific Support

Answered on Aug 30 2006

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