Hi Alice, Reasonable to exclude them from analysis. In a hypothesis-testing framework, you can ignore all the control probes, because we are not interested in their differential expression at all. If you choose to include them, it is an empirical way to empirical your false discovery rate, because we know that they should not show up. Simon ----- Original Message ----- From: "Johnstone, Alice" <alice.johnstone@esr.cri.nz> To: "Simon Lin" <simonlin at="" duke.edu="">; "bioconductor" <bioconductor at="" stat.math.ethz.ch=""> Sent: Sunday, March 09, 2008 8:56 PM Subject: RE: [BioC] What should happen to control probe information in beadlevel Illumina analyses? Thanks for your reply Simon, Yes the expression level is at the low end of the range so I do not think that this is a hybridisation problem as the original control plots looked fine. I used the VST transformation with a quantile normalisation of the Bead Summary data. My question is whether it is correct to remove the control probe information from the Bead Summary object before fitting the regression model so I don't get any spurious results like I have found, and what is the best way to do this? The expression values for one of the control genes are below, the trt1-control contrast comes up in my topTable results. This only occurs when I do my own bead-level summation, the BeadStudio produced bead summary data does not have the intensity info for this probe. Trt 1 Control Trt 2 7.544901 7.638846 7.531172 7.515447 7.614039 7.581452 7.495771 7.605357 7.555899 7.485725 7.622735 7.588895 7.538404 7.592215 7.516454 Regards Alice