In microarray experiments investigators sometimes wish to pool RNA samples before labeling and hybridization due to insufficient RNA from each individual sample or to reduce the number of arrays for the purpose of saving cost. The basic assumption of pooling is that the expression of an mRNA molecule in the pool is close to the average expression from individual samples. Recently, a method for studying the effect of pooling mRNA on statistical power in detecting differentially expressed genes between classes has been proposed, but the different sources of variation arising in microarray experiments were not distinguished. Another paper recently did take different sources of variation into account, but did not address power and sample size for class comparison. In this paper, we study the implication of pooling in detecting differential gene expression taking into account different sources of variation and check the basic assumption of pooling using data from both the cDNA and Affymetrix GeneChip microarray experiments.