Dear all, I have a question regarding duplicateCorrelation when applied to a paired design. I am analyzing 8 arrays (Agilent gene expression): 2 experimental groups A and B, samples are paired with one another: A1 with B1, A2 with B2 etc. Sample B2 has to be dropped because of quality issues, so one sample in group A missing its corresponding B. I have first processed an unpaired analysis of the data before processing a paired analysis (given the information I accessed later on). targets: FileName Group Pairs A1.txt A 1 A2.txt A 2 A3.txt A 3 A4.txt A 4 B1.txt B 1 B3.txt B 3 B4.txt B 4 ## Code design <- model.matrix(~0+targets$Group) colnames(design) <- unique(targets$ Group) # UNPAIRED ANALYSIS # fit <- lmFit(expr, design=design) contrast.matrix <- makeContrasts(contrasts="B-A", levels=design) fit <- contrasts.fit(fit, contrast.matrix) fit <- eBayes(fit) top.unpaired <- topTable(fit, coef="B-A", number=nrow(expr), sort.by="none") # PAIRED ANALYSIS # corfit <- duplicateCorrelation(expr, design, block=targets$Pairs) fit <- lmFit(expr, design=design, block = targets$Pairs, cor = corfit$consensus) contrast.matrix <- makeContrasts(contrasts="B-A", levels=design) fit <- contrasts.fit(fit, contrast.matrix) fit <- eBayes(fit) top.paired <- topTable(fit, coef="B-A", number=nrow(expr), sort.by="none") ## Results top.unpaired[1,] logFC AveExpr t P.Value adj.P.Val B 0.07913307 10.30572 0.7052201 0.49455794 0.9999397 -4.767979 top.paired[1,] logFC AveExpr t P.Value adj.P.Val B 0.08180912 10.30572 0.779451 0.451223001 0.9996952 -4.769007 What I do not understand is why the two logFC for the same probe are not similar? I can easily calculate logFC from the unpaired results, but not for the paired. I guess the fact that the paired design is "unbalanced" should be the reason; when I processed paired and unpaired analysis before removing sample B2 - 4 samples per experimental group - the logFC are identical. I would like to understand how it works: is there some weight applied to the samples in this case? I would also like to make sure that I don't make any mistake processing a paired analysis on this unbalanced paired design: should I also drop sample A2?? Any help and explanation is welcome. Thanks! Sarah