Optimization of oligonucleotide DNA microarrays is a multiparametric problem. The goal of the optimization process is to get conditions that capture target DNA with high sensitivity and selectivity. Parameters determining the performance of the microarray are spot morphology, probe and target density, background, and selectivity. More than 10 variables can be adjusted to obtain a well-optimized protocol. However, some variables only affect spot morphology and other factors affect for instance hybridization and selectivity, which can limit the optimization work considerably. This chapter suggests an outline on how an optimization procedure is made. Moreover, a simple method for absolutely quantify the number of hybridized target to the spots is given. The latter is important because it gives the possibilities to compare results with those obtained in the literature.
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