[BioC] About package beadarray

[Mark Dunning]

Hi Anand,

I think I understand the source of your confusion. There are two types
of correction applied to BeadArray data; one at the bead-level and one
at the summarized level.

The bead-level text files created by Illumina's scanning software
report the intensity of each bead on the array. These have been
adjusted by subtracting a local background measure calculated using
nearby pixels on the image. For convenience, the readIllumina function
in beadarray will read these values, as these are the same values that
are analysed by Illumina's BeadStudio/GenomeStudio software. We find
them adequate for most purposes, although a more thorough analysis is
possible as described in the imageProcessing vignette. If you have
data exported from BeadStudio they will almost certainly have been
adjusted in this way.

However, the backgroundNormalise method (that is now removed) referred
to in previous emails occurs on a per bead-type basis once data have
been summarized. It used to be the default normalization method in
BeadStudio. The quantities subtracted are negative controls
(bead-types with no target in the genome). As I mention in previous
posts there are viable alternatives to this method. The neqc method
you are interested in requires summarized data and (I suspect)
developed with GenomeStudio data in mind. Therefore it is Ok to use
data that have been read using readIllumina with this method.

Hope this helps. Regards,

Mark

On Thu, Jan 6, 2011 at 8:37 PM, Anand Patel <acpatel at gmail.com> wrote:
> Hmm.  This and the image processing file from the current dev version
> of the package (2.1.3) are seemingly inconsistent?
> http://bioconductor.org/packages/2.8/bioc/vignettes/beadarray/inst/doc/ImageProcessing.pdf
>
> Suggests that readIllumina always performs the backgroundNormalise process.
>
> http://bioconductor.org/packages/2.8/bioc/vignettes/beadarray/inst/doc/beadlevel.pdf
>
> and what's below suggests that this is not the case anymore.
>
> Could you clarify?  My goal in asking is to take data that's been read with:
>
> and use the neqc normalization process, which requires background
> correction not have been applied.
>
> Thanks,
> Anand
>
> On Tue, Dec 14, 2010 at 5:00 AM,  <bioconductor-request at r-project.org> wrote:
>> Message: 3
>> Date: Mon, 13 Dec 2010 14:24:45 +0000
>> From: Mark Dunning <mark.dunning at gmail.com>
>> To: Changhe Fu <fuchanghe at gmail.com>
>> Cc: bioconductor at r-project.org
>> Subject: Re: [BioC] About package beadarray
>> Message-ID:
>>        <AANLkTik5PWUwyhTBwWbqsoJcOCW9F68ED5uk8fEYfV=8 at mail.gmail.com>
>> Content-Type: text/plain; charset=ISO-8859-1
>>
>> Hi,
>>
>> That function used to be in beadarray to implement the background
>> normalisation that occurs in Illumina's BeadStudio/GenomeStudio.
>> However, it has been shown in various studies to be inferior to other
>> methods. See the neqc (limma) or vst (lumi) alternatives available
>> through the normaliseIllumina function in beadarray.
>>
>> Mark
>>
>>
>> On Fri, Dec 10, 2010 at 11:07 PM, Changhe Fu <fuchanghe at gmail.com> wrote:
>>> Hi, All
>>>
>>> In previous version of package beadarray, I can use the function
>>> backgroundNormalise(). But in new one, I cann't find it. Is it included in
>>> some other fuction of normalising?
>>>
>>> Thank you
>>>
>>> --
>>> Best,
>>> Changhe Fu
>>>
>>> ? ? ? ?[[alternative HTML version deleted]]
>>>
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>