Homemade Microarrays

Probe Labeling and Hybridization

The labeling method used will depend on the product being labeled. Most homemade arrays are designed to be used in competitive hybridizations, in which two probes are labeled with different colors and combined on the array. In general, the Bauer Core prefers using indirect labeling methods whenever possible. These methods involve incorporating an amino-allyl-labeled nucleotide into the product and subsequently coupling the dye to the amino allyl group (most often, Cy3 and Cy5 dyes are used). This method reduces dye bias, which may arise in direct labeling methods owing to differences in incorporation of each dye. Amino-allyl labeling takes a full day and the subsequent hybridization may be performed overnight. Each of the Bauer Core labeling protocols includes instructions for hybridization and washing. The reagent costs are about $30–40. Please direct questions about probe labeling to Christian Daly (cdaly@cgr.harvard.edu).

Amino allyl RNA labeling protocol (MS Word)
Genomic DNA labeling protocol (MS Word)