[BioC] Distribution of negative controls on Illumina HT12-v4 chips

[Wei Shi]

Hi Arnar:

	Although it is possible that the scanner upgrade could be the cause, but I think this possibility is quite weak. I will be happy to have a look at your control data to see if there is anything unusual with your controls. You can send me your control data only (the control probe profile) off-line if you like. I don't really think Illumina Technical Support can help you on this matter.

Cheers,
Wei

On Mar 1, 2011, at 5:42 PM, Arnar Flatberg wrote:

> Hi Gordon, Ina
> 
> 
> Thank you for your responses, they have been very helpful. We are currently
> seeking Illumina techsupport for answers. This might be related to a recent
> scanner upgrade.
> 
> 
> Arnar
> 
> 
> 
> On Mon, Feb 28, 2011 at 5:15 PM, Ina Hoeschele <inah at vbi.vt.edu> wrote:
> 
>> Hi Arnar, Gordon et al.,
>> we also have data on an increasing number of these chips,  and so far I
>> see no evidence for a bimodal distribution.
>> Ina
>> 
>> 
>> ----- Original Message -----
>> From: "Gordon K Smyth" <smyth at wehi.EDU.AU>
>> To: "arnar flatberg" <arnar.flatberg at gmail.com>
>> Cc: "Bioconductor mailing list" <bioconductor at r-project.org>
>> Sent: Saturday, February 26, 2011 11:39:20 PM
>> Subject: [BioC] Distribution of negative controls on Illumina HT12-v4 chips
>> 
>> Hi Arnar,
>> 
>> Wei Shi and I have checked out our own HT-12 v4 data, and we do not see
>> the bimodal distribution that you have observed.  So we think that this
>> might be a property of your data rather than a property of HT-12 v4
>> BeadChips in general.
>> 
>> Note that we are now recommending the neqc() function of the limma package
>> for background correcting and normalizing Illumina arrays.  This is a
>> modication of the normexp algorithm to use negative control probe
>> estimates.  See ?neqc and the Illumina case study in the limma User's
>> Guide.  This function will not be very sensitive to the shape of the
>> distribution of the negative controls.
>> 
>> Best wishes
>> Gordon
>> 
>> 
>> 
>> ------ ORIGINAL MESSAGE -------
>> [BioC] Distribution of negative controls on Illumina HT12-v4 chips
>> (double background?)
>> Arnar Flatberg arnar.flatberg at gmail.com
>> Tue Feb 22 15:46:45 CET 2011
>> 
>> Hi list,
>> 
>> We have recently seen an odd distribution of signal intensities using the
>> new version 4 HT-12 chips. Specifically, the distribution of negative
>> controls looks like a mixture of two gaussians. This invalidates the
>> common assumption of normal distributed background used in both the
>> detection calls and some error-model based normalizations, e.g. norm-exp
>> in limma. We did not have this type of distribution in version 3 of HT12,
>> nor in any other chip type from Illumina. Has anyone else seen this
>> particular issue, and specifically on the HT-12 v4 chip?
>> 
>> Below is a link to density plots of the negative control probes of 6
>> experiments I've compared. The top plots are from a lab in Oslo on the
>> HT-12 v4, at hte bottom are two experiments from our lab using HT12 v4 and
>> a comparison of a Rat v1 chip from our lab.
>> 
>> Figure:
>> http://oi52.tinypic.com/tz05v.jpg <%20http://oi52.tinypic.com/tz05v.jpg>
>> 
>> 
>> Thanks,
>> 
>> Arnar
>> 
>> 
>> ----------------------
>> Arnar Flatberg
>> Data scientist
>> Trondheim Genome Resource Center
>> St. Olav's hospital, Norwegian University of Science and Technology
>> Trondheim
>> Norway
>> 
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