I am analyzing in limma a two-color array experiment where I am compare 4 conditions: Wild-type (WT) versus Knock-out (KO) x Normal (NORM) versus Stress (STRESS). The hybs are replicated once (NNNNN_BLOCK1 and NNNNN_BLOCK2). I am most interested KOSTRESS-KONORM relationship for all genes on the array but would like to do all possible comparisons for my own reference. However, and I am sure this is due to a limitation of my intellect, I cannot figure out how to construct a design object to accomplish this. If the arrays shared a common reference (WT-ONLY), I would just do 'design <- modelMatrix (targets,ref="WT-ONLY")' but this is more complicated. Can someone please enlighten me? The content of the targets file for this experiment is at the end of this message. Thank you, Matt Vaughn FileName Cy5 Cy3 90930_BLOCK1.dat WTNORM KONORM 90930_BLOCK2.dat WTNORM KONORM 91003_BLOCK1.dat WTSTRESS KOSTRESS 91003_BLOCK2.dat WTSTRESS KOSTRESS -- Matthew W. Vaughn, Ph.D. Scientific Informatics Analyst Cold Spring Harbor Laboratory 1 Bungtown Road Cold Spring Harbor, NY 11724 phone: (516) 367-8469