chromatin IP troubleshooting

J Winkler news.100.johanneswinkler at spamgourmet.com
Thu May 4 16:16:31 EST 2006


the centrifuge speed for washing IP samples is critical.If the speed is 
too high, denatured protein can go down and mix with protein-A beads. 
with lower rpm, those denatured proteins should stick on side walls of 
eppis and you need to remove it from each wash step with a syringe-needle

Lynn Denekamp wrote:
> The ChIP assay has quite literally sucked the life out of me over the
> past year but I thought THIS TIME it would work. No such luck. My IgG
> negative control pulls down a product of the same intensity as the
> polII positive control antibody. Both antibodies came with the EZ ChIP
> kit (Santa Cruz).
> 
> The only advice I garnered from their protocol book was to decrease
> the number of PCR cycles and decrease the amount of DNA in the
> reaction. Any other suggestions?
> 
> Thanks.
> 


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