I've read capter 12 of the 2014 limma manual in order to identify DE genes in multiple arrays. There is one step where I'm not sure whether I did the right normalization.
In Cap. 12 the 'Aquantile' normalization between arrays is recommended:
> MA <- normalizeBetweenArrays(MA, method="Aquantile")
However, if I apply this to my 8-chamber Agilent data, I obtain a graph where the individual traces do not fit quite nicely.
I have found at Normalization for use of individual channels from 2-color arrays? that a combination of normalizeWithinArrays and normalizeBetweenArrays can be used instead. I used this:
MA.p <- normalizeWithinArrays(RG, method="loess")
MA.Aq <- normalizeBetweenArrays(MA.p, method="Aquantile")
plotDensitiesMA.Aq) now results in almost perfectly matching traces. Could anybody comment on this? Is this allowed?
Thanks for your replies!