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Highly Sensitive microRNA Array Performance

city was determined using probes designed with up to three mismatches. 10-fold more signal for perfect matches was obtained as compared to the same miRNA with <90% target homology. This experiment shows that individual miRNAs can be differentiated on arrays allowing detection of different miRNA isoforms that have three or more different nucleotides.


Precise Results

The simplest measure of precision for microarray analysis is self vs self analysis. For each tissue we split total RNA samples into two aliquots, and independently isolated miRNA from them. Each of the two aliquots was differentially labeled and co-hybridized to an miRNA array. Signal was measured for each element of the array. Since the differentially labeled samples were derived from the same total RNA sample, the two signals should have been approximately equal for each spot. We performed this experiment for a number of human samples; the average correlation of the data sets was routinely 9799% (Figure 3A).

Figure 3. miRNA Array Reproducibility. (A) Raw intensities from each channel (Cy3 and Cy5) for two self vs. self arrays. The arrays were performed on two separate days. The average correlation of such samples routinely ranges between 9799% indicating precision with the array system. (B) The average Log2(R/G Normalized Ratio) of each spot on the array from six independent prostate vs colon samples is plotted as a circle on the graph. The average standard deviation of each spot is indicated by the error bars. The average correlation for the 6 replicates is 98% indicating reproducibility with the array system.
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