During WB, a huge background is caused by IgG. Are there any methods by which I can reduced this background?
I use Dynabeads that are linked with anti IgG. Its like to precipitate/pull down all the IgG in the lysate.
http://products.invitrogen.com/ivgn/product/11201D
Hi, try protein A of protein G beads (Pierce) to clear IgG from you serum samples. If you have a lot of material you could consider using HiTrap columns (GE Healthcare). The nice thing is that you can reuse these beads or columns several times without efficiency loss and as such they probably provide a cheaper alternative that the Dynabeads.
IgG can be bound to ProA/G beads, by just passing the plasma over the column or, incubate the beads with the plasma for 1 hr and centrifige the beads dowm, again u have to check whether u are not loosing the protein of interest on the PrA/G beads nonspecifically. Swell some PrA/G beads in 1xPBS for 1 hr and u can use them directly to the plasma. secondly, it is a crude way but if not effects the protein of interest u can try that too. 0.2gm of Ammonium sulfate per ml selectively precipitates the total igG in the plasma. After 1 hr of incubation at 4 deg with AASo4, spin it outt and the plasma can be loaded (if needed dialysis is additional step). The second method is time taking and crude too, and u have to standardize for your protein
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