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Reference protein give different expression

Western blot

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3 replies to this topic

#1 tierarztkarakus

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Posted 17 February 2013 - 03:35 PM

Hi all, I've been problem with beta-actin. The actin bands give different expression each other sample (between control and other treated sample). Picture attached. Help please.
What can i do?
Has anyone suggestion



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#2 jerryshelly1

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Posted 17 February 2013 - 04:59 PM

Do a coomaisse and look at your relative loaded protein concentrations.

#3 Tabaluga

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Posted 17 February 2013 - 06:27 PM

The actin bands give different expression each other sample (between control and other treated sample).


Didn't quite understand that, are the actin bands always stronger or weaker depending on whether the cells were treated ot not ? If so, it could be that your treatment influences actin levels and I'd try another loading control (GAPDH, lamin B1....)
If not, you should determine the load with a Coomassie as indicated in the post above.

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La chose simplement d'elle-même arriva,
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#4 bob1

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Posted 18 February 2013 - 12:04 AM

Or determine the total protein concentration beforehand (e.g. bradford or BCA assays) and then load based on that.





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