I have seen this question posed elsewhere, but wanted to double check for some of my colleagues who want to make sure. It is the following: when defining contrasts in Limma, how is the syntax to be interpreted? I've included some code. If clarification is required, I can provide that as needed.
#### edata is expression matrix of aptamer probes; design is the design matrix; Used a repeated measures design so estimated the patient-wise correlation in expression over visits where StudyID defines this pairing ; pheno_table details sample metadata fit<-lmFit(edata,design, block=pheno_table$StudyID, correlation=corfit$consensus) cm <- makeContrasts( `Diabetpost.PUFA.Met-Diabetpre.PUFA.Met` = Diabetpost.PUFA.Met-Diabetpre.PUFA.Met, `Diabetpost.MUFA.Met-Diabetpre.MUFA.Met` = Diabetpost.MUFA.Met-Diabetpre.MUFA.Met, `Diabetpost.PUFA.NoMet-Diabetpre.PUFA.NoMet` = Diabetpost.PUFA.NoMet-Diabetpre.PUFA.NoMet, `Diabetpost.MUFA.NoMet-Diabetpre.MUFA.NoMet` = Diabetpost.MUFA.NoMet-Diabetpre.MUFA.NoMet, levels=design) fit2 <- contrasts.fit(fit, cm) fit2 <- eBayes(fit2,robust=TRUE) T<-topTable(fit2,number=100000)
In plain English, are the above comparisons made "between post with respect to pre" or "between pre with respect to post"? That is, would a positive logFC indicate post > pre or pre > post? Again, I believe I know the answer, but I would like confirmation for my colleagues. Thanks.