Gene expression analysis using cDNA microarrays

This unit focuses on cDNA microarrays, which are being increasingly applied to large-scale gene expression studies in the nervous system, concentrating on the steps of probe preparation and microarray hybridization. Four protocols for the attachment of fluorophores to cDNA (labeling) and cDNA hybridization onto microarrays are presented: direct labeling using either Klenow fragment or reverse transcriptase, and two signal amplification techniques represented by the tyramide signal amplification method (TSA), and detection and labeling of cDNA using PCR amplification. Basic issues in experimental design, sources of variability and potential pitfalls in microarray experiments are described. Anticipated results and follow up of microarray experiments, including confirmation of the results and troubleshooting are also discussed.