Question (33714) | PC-12 nuclear extract lysate (ab14884)

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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.

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.