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I was trying to find on web more data for that Ab searching by clone name (Jol5). I did miss to mention that this data are not on Abcam website and this caused your confusion. Sorry for that. There are actually several manufacturers of this Anti-Lamin A/C Ab that used (clone Jol5). I assumed that same clone will give the same Ab with the same results on WB. The fact that made me concerned about the specificity of ab5090 was that anti-desmin Ab D1030 (Sigma) recognized same molecular weight band (~53kD-correct for desmin) as ab5090 did. I have to mention that the conjugate I used has a background signal in the same MW area, but it is less intense than the positive signal. In the mean time I tried anti-laminA Ab from Chemicon with another conjugate (without background signal) and received a signal corresponding to ~70kD for lamin A. In all experiments I used same protein sample and MW-marker and % PAA gel, so I do not expect that Lamin A/C proteins would migrate differently in all blots. To summarize all, antibody ab5090 that I tested at dilution 1/50 for detection of Lamin A/C in protein extracts from heart tissue showed a weak positive signal of MW ~53kD, that is smaller than expected molecular weights for Lamin A/C. I hope this will answer your questions. Thank you for your help. I'll be happy if you can recommend me a protocol for immunostaining with ab5090 antibody.
Asked on Nov 19 2004
My sincere apologies for the delay in answering your enquiry, I have heard back from the source of Jol5 who has provided me with this protocol: To perform IF on C2C12 myoblasts with JoL5 we fix with 1:1 methanol:acetone. The methanol:acetone is prechilled to -20C and the cells are subsequently fixed for 10 minutes at 4oC. The staining is predominantly observed in post-mitotic myoblasts and myotubes. For blotting it also works nicely with standard protocols. As mentioned before JoL5 recognises a growth specific epitope. It is expressed in non dividing cells but is undetectable in growing cells. HeLa is a growing cell. You will need to use ab4789 which detects lamin A/C in all cells. I hope this information helps,
Answered on Dec 08 2004