miChip: A microarray platform for expression profiling of microRNAs based on locked nucleic acid (LNA) oligonucleotide capture probes

As key regulators of post-transcriptional gene expression, it is important to monitor the expression of microRNAs (miRNA) in diverse physiological and pathophysiological processes. Here, we describe a method for sensitive and accurate microarray-based expression profiling of miRNAs. The protocol focuses on the use of locked nucleic acid (LNA)-modified capture probes. LNAs are bicyclic nucleotide analogues that significantly increase the melting temperature (T.) of hybrids with miRNAs. Mixed LNA/DNA capture probes thus can be designed for equal T(m)s for all miRNAs, which naturally cover a range between 45 and 74 degrees C. The protocols established are easy to apply, as they do not require RNA size selection and/or amplification of miRNAs. Moreover, they enable high affinity hybridizations yielding accurate signals that discriminate between single nucleotide differences and hence closely related miRNA family members. (C) 2007 Elsevier Inc. All rights reserved.