Heavy chain "contamination" with Ip - (Jun/12/2007 )
Hi,
I need to understand a really basic problem. I am doing an Ip with an antibody that is raised in mouse. After, I do the WB with an antobody from rabbit to do not have the heavy chain contamination band of the first Ip antibody. But still, I need to do a loading control and for that I need to blot with the first antibody that is raised in mouse, but I will automatically have the contamination band that is close to my protein size. Do you have a solution different than try to find and buy an antibody that is made from another species?
Thank you very much for your help!
Brassap
-brassap-
QUOTE (brassap @ Jun 12 2007, 10:42 PM)
Hi,
I need to understand a really basic problem. I am doing an Ip with an antibody that is raised in mouse. After, I do the WB with an antobody from rabbit to do not have the heavy chain contamination band of the first Ip antibody. But still, I need to do a loading control and for that I need to blot with the first antibody that is raised in mouse, but I will automatically have the contamination band that is close to my protein size. Do you have a solution different than try to find and buy an antibody that is made from another species?
Thank you very much for your help!
Brassap
I need to understand a really basic problem. I am doing an Ip with an antibody that is raised in mouse. After, I do the WB with an antobody from rabbit to do not have the heavy chain contamination band of the first Ip antibody. But still, I need to do a loading control and for that I need to blot with the first antibody that is raised in mouse, but I will automatically have the contamination band that is close to my protein size. Do you have a solution different than try to find and buy an antibody that is made from another species?
Thank you very much for your help!
Brassap
How about do it the other way around, IP with rabbit Ab and WB with mouse Ab?
-Almasy-
QUOTE (Almasy @ Jun 18 2007, 06:42 AM)
QUOTE (brassap @ Jun 12 2007, 10:42 PM)
Hi,
I need to understand a really basic problem. I am doing an Ip with an antibody that is raised in mouse. After, I do the WB with an antobody from rabbit to do not have the heavy chain contamination band of the first Ip antibody. But still, I need to do a loading control and for that I need to blot with the first antibody that is raised in mouse, but I will automatically have the contamination band that is close to my protein size. Do you have a solution different than try to find and buy an antibody that is made from another species?
Thank you very much for your help!
Brassap
I need to understand a really basic problem. I am doing an Ip with an antibody that is raised in mouse. After, I do the WB with an antobody from rabbit to do not have the heavy chain contamination band of the first Ip antibody. But still, I need to do a loading control and for that I need to blot with the first antibody that is raised in mouse, but I will automatically have the contamination band that is close to my protein size. Do you have a solution different than try to find and buy an antibody that is made from another species?
Thank you very much for your help!
Brassap
How about do it the other way around, IP with rabbit Ab and WB with mouse Ab?
That's exactly what I did. I have a monoclonal antibody that bind a protein. After, I have to do my WB with an antibody (rabbit) that bind only the phosphorylated form of that protein. Then, I have to verify the loading with the total protein but again, this is with the same monoclonal antibody than the IP, so that is why I have the contamination band. But I found an antibody agains the light chains, I will try...
Thanks!
Brassap
-brassap-