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Loading control for serum samples?

Loading control for serum samples? - Western Blot Forum

Loading control for serum samples? - Discuss western blotting and immunoblot in the western blot forum discussion board. Ask Questions about transfers, blocking, membrane antibody incubation and exposures.


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Old 01-07-2009, 04:30 PM
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Default Loading control for serum samples?



I'm comparing large numbers of dog serum samples by W. blotting. I'm wondering what a good loading control is for this sample type, albumin was my first thought but it's not at all in the same order of magnitude as my signal, and it seems to vary in different sera.

Does anyone have any recommendations?
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Old 02-27-2009, 02:29 PM
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Default Re: Loading control for serum samples?

BUMP

Anybody?
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Old 02-27-2009, 02:52 PM
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Default Re: Loading control for serum samples?

Hello Allison,
welcome to the forums

have you tried tubulin, actin or other like albumin? I am not sure however what your treatment affects, do you see any changes in such proteins?
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Old 03-17-2009, 11:27 PM
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Default Re: Loading control for serum samples?

Hello Allison,

you can test an enzyme at 10 kDa. and be carful that this enzyme will not affect your analyse.

good luck
ayman
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Old 03-18-2009, 01:04 PM
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Default Re: Loading control for serum samples?

That's an interesting reply... which enzyme at 10 kDa... I expect there are quite a few

(anyway I'm trying actin, b-tubulin, and eventually GAPDH. I will post results, if any, here)
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Old 03-24-2009, 05:27 AM
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Default Re: Loading control for serum samples?

Hey Allison,

I'm trying to run a WB with some rat plasma samples. I wouldn't recommend using GAPDH... didn't work for me. I'll let you know if I find anything that works... and I'd appreciate it if you could return the favour.
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Old 04-17-2009, 05:50 PM
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Default Re: Loading control for serum samples?

I might try using Ponceau S - which stains proteins generally - as a loading control. It seems somewhat sketchy, but others have reported doing so in the literature, presumably when standard loading controls were unreliable.

See Olesen & Auger, 2005, J Neuroendocrinol, 17, 255-261 for an example... I'd post the DOI link, but apparently I need to prove that I'm not spamming the forum first...

Good luck.
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