Effect of target length on specificity and sensitivity of oligonucleotide microarrays: A comparison between dendrimer and modified PCR based labelling methods

• Target length should be tailored to increase specificity without compromising sensitivity.
• PCR labelling is superior to the dendrimer approach in maximizing hybridization efficiency.
• Replacing FAST Frame by Hybrislip for hybridization increases sensitivity by 100 fold.
• Denatured samples kept at room temperature produces higher sensitivity than those kept at hybridization temperature.

DNA microarrays are widely used as end point detectors for gene expression analysis. Several methods have been developed for target labelling to enable quantification but without taking target length into consideration. Here we highlight the importance of choosing the optimum target length that would ensure specificity without compromising sensitivity of the assay. For this, eight plasmids that are identical to each other except for a closely related 23 bp unique reporter (UR) sequence were used to examine the hybridization efficiency for these URs. Targets of various lengths were generated and labelled as follows: full length and 330 bases transcripts using a dendrimer labelling method, 120 bp amplicons by the modified PCR end labelling method and synthetic labelled targets of 33 bases. This report also shows the advantages of using the modified PCR method over other labelling methods in generating labelled amplicons of the desired lengths to maximize hybridization efficiency.