Hi there, I am designing a pilot microarray study on embryoic developmental mutant using affy platform. The comparison itself is very simple, the mutant vs normal at one time point. Due to various reasons (mostly funding and limited amount of tissue), I can't start with the "ideal" approach in which each sample is hybridized to an individual chip. Since I can easily rear a lot of animals, it seems that pooling is the only choice for the pilot study. However I am not sure what is the best way to allocate the pooled samples to each chip. For example if I want to do 3 array replicates each for the mutant and control. Is it better to pool enough samples for 3 arrays and then separate the pooled sample in 3 portions for hybridization or just pool different individual samples for different replicates? It seems to me that the first way is like getting a group expression average with accessment of technical variation, while the second approach can also provide some sort of evalution of biological variation, abeit an averaged one by the pooling. I suspect the latter approach is better, and would love to know the suggestions from you. Thanks! Fai -- Yuk Fai Leung Department of Molecular and Cellular Biology Harvard University BL 2079, 16 Divinity Avenue Cambridge, MA 02138 Tel: 617-495-2599 Fax: 617-496-3321 email: yfleung@mcb.harvard.edu; yfleung@genomicshome.com URL: http://genomicshome.com