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Getting over 100% Input in IP'd samples

Getting over 100% Input in IP'd samples - ChIP Chromatin Immunoprecipitation Forum

Getting over 100% Input in IP'd samples - Chromatin Immunoprecipitation Forum. Discuss, chat about, and post questions about the ChIP assay, in the ChIP forum. Includes troubleshooting, protocols, buffer preparation, and more.


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Old 09-09-2010, 05:26 AM
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Default Getting over 100% Input in IP'd samples



Hi,

I've been doing ChIP for the first time and using an antibody for histone H3 trimethyl K27, and Mouse IgG for negative control. my results were looking good, and someone advised me to also look at an antibody for pan Histone H3 as a positive control.

The problem is that in some of my samples, and only in some of my primer sets, my H3 immunoprecipitated samples are returning over 100% input - which is, of course, ridiculous. I even got over 500% in one sample. Other primer sets are returning more sensible numbers - like 10-20%.

I fear that this may indicate a problem with either the IP protocol in general, or a problem with the handling of input controls. I am using the Invitrogen MAGnify kit for immunoprecipitations.

Has anyone experienced a similar strange result?
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% input , 100% , chip , histone h3 , input , ipd , samples


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