PC-12 nuclear extract lysate (ab14884)

Submit a review Q&A (4)

Overview

  • Product name

    PC-12 nuclear extract lysate
    See all PC12 lysates

  • General notes

    Extracts have been quality control tested by Western blot and the Electrophoretic Mobility Shift Assay (EMSA). 

Properties

  • Mycoplasma free

    Yes

  • Form

    Liquid

  • Storage instructions

    Shipped on dry ice. Upon delivery aliquot and store at -80ºC. Avoid freeze / thaw cycles.

  • Storage buffer

    pH: 7.90
    Constituents: 0.0002% Sodium molybdate, 0.216% Beta glycerophosphate, 0.018% Sodium orthovanadate, 0.042% Sodium fluoride, 0.015% DTT, 0.48% HEPES, 0.003% EDTA, 20% Glycerol, 2.34% Sodium chloride

    With Protease inhibitors
  • Concentration information loading...

  • Research areas

  • Background

    Generated by Greene and Tischler (1976) from a transplantable rat adrenal pheochromocytoma line, PC12 cells respond reversibly to nerve growth factor (NGF) and therefore have a neuronal phenotype, making the line a useful model system for neuronal differentiation. They produce catecholamines (dopamine, norepinephrine), neurotransmitters, NPP and are therefore used in studies of differentiation, neurobiochemistry and neurobiology.

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

References (0)

Publishing research using ab14884? Please let us know so that we can cite the reference in this datasheet.

ab14884 has not yet been referenced specifically in any publications.

Customer reviews and Q&As

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1-4 of 4 Abreviews or Q&A

Question

Thank you for your prompt reply. Please send the HeLa nuclear lysate that you described; just to confirm it is a nuclear lysate and not a cell lysate? I appreciate your help and patience with this matter.

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Answer

Thank you for your reply.

The product ab29545 is a whole cell lysate not a nuclear lysate. The only nuclear lysate we have in stock is the ab14655that you currently have.

If you still wish for me to send you ab29545, the whole cell lysate, please let me know and I will have it sent today. Otherwise, I can refund the cost of ab14655, as it did not produce a signal for you.

I look forward to your reply and resolving this issue for you.

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Question

I must have misunderstood you, I loaded 18uL per well (I had 10uL of HeLa nuclear lysate and added 10uL of 2x protein loading buffer to a total volume of 20ul of which I loaded 18ul per well). The wells themselves if loaded carefully can hold 22uL of sample.

Do you recommend I try to load 4uL more?

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Answer

Thank you for that information.

I do not think adding 4ul more will increase your signal and since you can only load a maximum of 22ul/well, I would like to send you ab29545 as a free of charge replacement. This HeLa cell lysate is already in sample buffer and is at a higher concentration that the original one sent to you, so you can load more total protein and hopefully see a signal with your 3 antibodies. If this is ok with you, please let me know and I will process the replacement for you.

I look forward to you reply.

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Question

I loaded 2.5mg/ml of total protein (the original concentration) I also tried lower concentrations (0.5 mg/mL and 0.25 mg/mL) and saw signal only in the PC12 nuclear lysate of corresponding concentrations. I assume that if the PC12 lysate showed a clean signal at 2.5mg/mL the HeLa lysate should also work - but it did not.

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Answer

Thank you for your reply.

I appreciate your patience in this matter, but when you say you loaded 2.5mg/ml of total protein, how many microlitres of lysate is that? I ask, as we send 100ug of total protein at at concentration of 2.5mg/ml (therefore we you should have received 40ul total volume). I just want to make sure that the problem is with the lysate and not that more needs to be used. Although you used both the HeLa and PC12 lysates at the same concentrations, it may be that you need to use more of the HeLa lysate, as you were seeing a signal for Tubulin (the signal was weak, but it was present).

Once again thank you for your patience in this matter, if it does turn out that there is a fault with the lysate, then I would be more than happy to send a new vial.

I look forward to your response.

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Question

On January 10, 2012 I placed an order for HeLa and PC12 nuclear lysates (order confirmation # **********). I ran both lysates at 2.5 mg/ml (original concentration) and stained the blot with three different antibodies; among them with a anti-TATA binding protein antibody raised against a peptide mapping at the N-terminus of TBP of human origin. The species reactivity as specified by the manufacturer of the anti-TBP was to human, rat and mouse lysates - the antibody worked for the PC12 nuclear lysate (ab14884) but not for the HeLa nuclear lysate (ab14655). Likewise the two other antibody stains worked for the PC12 nuclear lysate but not for the HeLa nuclear lysate (anti-NSF and anti-tubulin).

This makes me believe that perhaps the HeLa nuclear lysate was in some way faulty and I was wondering if you may offer me some advice.

Please see the attached blot for further details.

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Answer

Thank you for contacting Abcam.

I am sorry about the problems you have been having with ab14655, HeLa cell lysate. I was wondering if you could tell me how much total protein you loaded of the cell lysate. I ask as there does seem to be a signal (a weak signal) for Tubulin and I am wondering if you load more of the lysate onto the gel, you would increase the signal for Tubulin and also begin to see a signal for the other antibodies.

If increasing the total amount of protein in the well does not improve the signal, then I would be happy to send you another aliquot of the HeLa lysate.

I look forward to your reply and helping you resolve this issue.

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