When you performed the Co-IP, did you add loading buffer and boil the samples? It may be that you are detecting your protein of interest and the antibody (which is another protein). Sounds like you may need to wash better. An image of your staining would be helpful. I need to know what the MW of your TOI is and then based on the image with MW markers, I can have a better idea of what is going on.

Are the two bands at 25 and 50kDa? These are the size of the light and heavy chains of the antibody (which you most likely still have some coming off your beads).

Hi there,

Haven't you got an other detection mode than Coomassie staining? As William said you are certainly seeing Ig light and heavy chains on gel. I think you need something more sensitive to check exactly what is the content of your IP (ie WB with a specific antibody).

I agree with William Grey's comment, it might be heavy and light chain. If you can add picture then it will be easy to analyze. Why dont you go for silver staining? as concentration of Co-IP sample is less and silver staining is more sensitive to detect proteins at lower concentration.

I think silver staining or WB will show your protein of interest. If the bands you are seeing are at unspecific MW stringent washing steps could help. 

Thanks for the suggestions. I have attached the pic of the gel. The first lane is the control, second and third are the test. I did heat the samples with the sample buffer as I do for western. When i ran the sample on western and probed with CDMP1 antibody which is my protein of interest, I do still bands corresponding to the bands that I see on the gel. So what I thought was these are the non specific bands of the antibody. Please add your comments.

I guess these are not the heavy or or light chain bands. These are might be non specific bindings. 

Why dont you load supernatent?

Why dont you boil? If you dont boil then then how come proteins get eluted/seperated from beads?

Best way to go for silver staining

As you said I think these are the non specific bands, but the primary ab is poly clonal and it binds non specifically to many proteins. Is there any way to minimize this

What do you use to block your membrane and dilute your antibody into? Blocking with milk helps to prevent non-specific binding. Depending on where the antibodies are from, you can also use milk, to reduce non-specific band formation. Always use secondary antibodies in milk. 

I agree with Anne von Koschembahr, You may try skim milk (1, 3 or 5%) if it will not work then you can go for BSA (1, 3 or 5%) for blocking.

There was no difference when I used milk or BSA for blocking. I still see the same bands on western