2 replies to this topic

[ken_zhang]

I am doing the western blot.there is no problems when I transfer the proteins to PVDF menbrane.But I got a high background.

[protolder]

ken_zhang, on 21 April 2011 - 11:25 PM, said:

I am doing the western blot.there is no problems when I transfer the proteins to PVDF menbrane.But I got a high background.

Hola, change the blocker agent or block for more time, because if you have background is because primary ab. bounds lightly but inespecifically to the membrane,I think. Buena suerte

[steffi333]

ken_zhang, on 21 April 2011 - 11:25 PM, said:

I am doing the western blot.there is no problems when I transfer the proteins to PVDF menbrane.But I got a high background.

Try blocking with 5% non-fat milk in PBS. Also dilute your antibodies in this. Also I've found that if you use your primary at too high a concentration you can get a lot of background. And wash LOADS! With plenty of wash buffer.