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I've been thinking about this and the only thing I can possibly think of is that during the DNase treatment, I add DNase inactivation reagent, mix it, then centrifuge. I'm using the kit from Ambion. After centrifugation, the inactivation reagent forms a white "pellet" at the bottom. But I've noticed that it's quite "loose" and that it is disturbed quite easily. I try to avoid carryover, but there isn't any guarantee as it is quite difficult to tell.
I've been told you don't want carry over as this could inhibit other enzymes etc. If some of my biological repeats have this DNase inactivation reagent as contamination, I'm guessing this could lead to variability with my biological repeats?
Any other suggestions?
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In Topic: Trouble with microarray. Too much variability between biological repeats
14 September 2012 - 08:46 AM
pcrman, on 14 September 2012 - 07:54 AM, said:
You did not mention technical repeats. Have you included them, if yes, what is the variability between them? Without first knowing variability between technical repeats, it is hard to see what caused the variation between biological repeats.
Technical repeats weren't included, if there was variability between technical repeats, then I assume the problem is coming from the technicians end. But she has done microarrays on swarm cells before that have worked, so I think the problem is coming on my end. I just have no idea what is causing it since my part is so straight forward.
