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Inaccuracies in protein MW on SDS-PAGE

Inaccuracies in protein MW on SDS-PAGE - SDS-PAGE Gel Electrophoresis Forum

Inaccuracies in protein MW on SDS-PAGE - SDS-PAGE Forum and Gel Electrophoresis Forum. Discuss the running of agarose gels, sds-page gels, and other gels including sequencing, gradient or tricine.


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Old 02-28-2010, 01:58 PM
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Default Inaccuracies in protein MW on SDS-PAGE



Hello everyone, got a quick question about the reliability of MW estimations using SDS-PAGE. I've got a protein which is estimated to be ~60 kDa, although on my gel it seems to consistently migrate closer to 65 kDa.

I know that SDS-PAGE may not always be accurate for measuring these things, but is there any way to improve on this? What is the reason for the higher migration? Improper denaturation?

Thanks alot.

S
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Old 04-22-2010, 12:16 PM
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Default Re: Inaccuracies in protein MW on SDS-PAGE

There are a lot of possible explanations.
First of all are you using a pre-stained marker?. Pre-stained marker gives an approximate idea of the molecular but unstained markers are more precise. May be you could run BSA (Molecular weight: 66382) in one well to check if your marker is behaving good.
Other options are:
-your protein may have post-traductional modifications (glicosilation, phosphorilation, an so on)
-your loading buffer could be interfering with your run.
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