more protein can give a band shift on a SDS-PAGE? - (Nov/13/2007 )
I explain my problem.... I run an SDS-PAGE and blot it in order to do a Western blot, samples were IgG and the primary Ab was anti-Fc.
The results: I found lot of bands at 150-125-100-75-50 KDa (not good for my protein ) in NON REDUCING CONDITIONS: so I supposed that all this stuff could be partial molecules (2heavy chain + 2 light chain, 2 heavy chain + 1 light chain, 2 heavy chains only, 1 heavy chain).
When I loaded reduced samples my suppositions were quite confirmed: I found only ONE band at ~55 KDa (heavy chain, I suppose). The main problem is that this band is a little higher than the 50 KDa band that I found in non reducing conditions.
Here my question: is it possible that since in reducing conditions all the bands become one (sorry for the bad expression ) this band migrates slowly than the other 50 KDa non reduced ones?
The results: I found lot of bands at 150-125-100-75-50 KDa (not good for my protein

When I loaded reduced samples my suppositions were quite confirmed: I found only ONE band at ~55 KDa (heavy chain, I suppose). The main problem is that this band is a little higher than the 50 KDa band that I found in non reducing conditions.
Here my question: is it possible that since in reducing conditions all the bands become one (sorry for the bad expression

Can you give us a picture of your gels (reduced vs non-reduced)? That will help immensely.
Typically, when you overload a gel, the band's leading edge tends to stay at the correct size, and the band just gets thicker; it may tend to broaden the lane, so that adjacent lanes get pushed aside (this might be the best indication of overloading). If you're not sure about this, try loading less and see what effect this has.
As for the apparent difference in size, your non-reduced band is going to have a slightly more compact shape, so it will tend to run a bit faster. I wouldn't worry; the fact that both bands appear by Western means that they are the same thing, so the size difference has to be due to the sample preparation (reduced vs non-reduced). Just remember tht you should be able to explain the difference in your paper/thesis/report.