CHIP Assay

UchiNews anybody at uchicago.edu
Tue Mar 18 23:22:57 EST 2003


Seems like you proteins bind to the beads non-specifically and carry over
DNA bound to them.
You could try:
1) playing with salt concentration in your buffer (0.5-1M NaCl)
2) different buffer pHs - you might want to select pH that is low enough to
reduce non-specific binding while still allowing Ab/Ag interaction.

If the proteins you work with bind specific DNA sequence, could adding
non-specific DNA (e.g. a 50-100 mer of non-coding satellite DNA) help reduce
false positives?

-Emir


<ebodzak at ucalgary.ca> wrote in message
news:20030318231228.14262.qmail at ww02.hostica.com...
>
> Does anyone out there have any suggestions on how to alter the typical
CHIP protocal to avoid false positive results? During the assay a specific
antibody is added to bring down protein-DNA complexes, and the DNA is then
examined by PCR. In order to ensure the DNA being brought down is specific,
a negative control is used, a non-IP'd sample. Without the addition of
antibody no complexes should be coming down, but in my case DNA is coming
down! My negative control is positive! Does anyone know how to adjust the
protocal to avoid the false positives in the negative control?
>
>
http://biowww.net/mynews/tree.php?group_name=bionet_molbio_methds-reagnts&be
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