[BioC] silly question on how to use arrayMagic on agilent two-color arrays

[Sean Davis]

James Anderson wrote:
> Hi, 
>
> I am analyzing about 30 agilent two color arrays. One simple question is that, in terms of the downstream analysis such as finding differentially expressed genes, what are the features should I use (such as log ratio, mean intensity of red and green, etc)? I have some experience in Affy, but almost no experience in agilent.
>
> In addition, I am trying to use arrayMagic from Bioconductor to do quality control and pre-processing of my agilent arrays. From the example arrayMagic gave in the manual, it seems that the format of the example data is somewhat different than agilent data format. 
>
> I saw the following header file 
>
>        HEADER       SPOT       GRID       TOP       LEFT       BOT       RIGHT       ROW       COL       CH1I       CH1B       CH1AB       CH2I       CH2B       CH2AB       SPIX       BGPIX       EDGE       RAT2       MRAT       REGR       CORR       LFRAT       CH1GTB1       CH2GTB1       CH1GTB2       CH2GTB2       CH1EDGEA       CH2EDGEA       FLAG       CH1KSD       CH1KSP       CH2KSD       CH2KSP   
> While my agilent arrays has the following header information:
>
>        FeatureNum       Row       Col       Start       Sequence       ProbeUID       ControlType       ProbeName       GeneName       PositionX       PositionY       LogRatio       LogRatioError       PValueLogRatio       gSurrogateUsed       rSurrogateUsed       gIsFound       rIsFound       gProcessedSignal       rProcessedSignal       gProcessedSigError       rProcessedSigError       gNumPixOLHi       rNumPixOLHi       gNumPixOLLo       rNumPixOLLo       gNumPix       rNumPix       gMeanSignal       rMeanSignal       gMedianSignal       rMedianSignal       gPixSDev       rPixSDev       gBGNumPix       rBGNumPix       gBGMeanSignal       rBGMeanSignal       gBGMedianSignal       rBGMedianSignal       gBGPixSDev       rBGPixSDev       gNumSatPix       rNumSatPix       gIsSaturated       rIsSaturated       PixCorrelation       BGPixCorrelation       gIsFeatNonUnifOL       rIsFeatNonUnifOL       gIsBGNonUnifOL       rIsBGNonUnifOL       gIsFeatPopnOL       rIsFeatPopnOL    
>    gIsBGPopnOL       rIsBGPopnOL       IsManualFlag       gBGSubSignal       rBGSubSignal       gBGSubSigError       rBGSubSigError       BGSubSigCorrelation       gIsPosAndSignif       rIsPosAndSignif       gPValFeatEqBG       rPValFeatEqBG       gNumBGUsed       rNumBGUsed       gIsWellAboveBG       rIsWellAboveBG       gBGUsed       rBGUsed       gBGSDUsed       rBGSDUsed       IsNormalization       gDyeNormSignal       rDyeNormSignal       gDyeNormError       rDyeNormError       DyeNormCorrelation       ErrorModel       xDev       gSpatialDetrendIsInFilteredSet       rSpatialDetrendIsInFilteredSet       gSpatialDetrendSurfaceValue       rSpatialDetrendSurfaceValue   
>
> How should I use arrayMagic on my data?
>   
I haven't used arrayMagic for Agilent data.  You can use limma and 
read.maimages() to load your data.  As for what columns to use, the 
logRatio column of course contains a ratio, while the columns 
rProcessedSignal and gProcessedSignal probably contain the 
background-corrected intensities.  You will probably want to glance at 
the Agilent technical manual about what the columns represent, in more 
detail.  As for what columns you will want to use in your analysis, you 
will need to be specific about your experimental design.  If you used a 
common reference, then you can simply treat the logRatio in the same way 
as your affymetrix arrays for the purposes of analysis.

Sean