The Genomics and Microarray Laboratory (GML) was set up as a core facility for investigators at the University of Cincinnati and UC Medical Center to provide services at cost.  The GML has a 48-pin microarrayer, two dual laser fluorescent scanners, an Agilent Bioanalyzer, a tetrad thermocycler for 96- and 384-well plates, and a robotic machine dedicated to high-throughput liquid dispensing. 

The microarrayer has the capacity to spot approximately 60,000 DNA samples per slide, and the scanner reads Cy3- and Cy5-fluorescent tagged, target cDNA. 

The spotted DNA sample (known sequence) is referred to as the probe, and the fluorescent-labeled cDNA (unknown sequence) is referred to as the target.

Two RNA samples for target labeling, a control and experimental, are used per slide.  A minimum amount of 20mg total RNA or 0.5mg poly(A)+ RNA is recommended for each RNA sample.  Lesser amounts of RNA are acceptable, and for very small amounts of RNA when RNA is difficult to obtain, an amplification/labeling protocol is used.  The RNA provides the template to generate fluorescent-labeled (cytidine-3 and cytidine-5) target cDNA.   Hybridizations and washes are carried out as with conventional DNA blotted filters.  The slides are scanned and differential gene expression levels are determined by the calculated ratio of Cy-3 to Cy-5 emittance.

To determine experimental variability, it is strongly recommended that a given experiment be carried out at least in triplicate.  For the same reason, it is also strongly recommended that for one of the slides in the triplicate, the Cy-3 and Cy-5 labeling be switched for the control and experimental target RNAs.