RNA Amplification

Typically our microarray hybridization experiments require 1-2 ug of total RNA. However, many investigators are performing experiments using small amounts of material (e.g. sorted cells, laser capture or islets). We can successfully amplify RNA from as little as 5-25 ng of total RNA (1000 cells or 1 islets). We use Nugen’s WT-Ovation™ Pico or Ovation V2 RNA Amplification and BioPrime Labeling System which involves a series of enzymatic reactions resulting in linear amplification of exceedingly small amounts of RNA for use in array analysis. Unlike exponential RNA amplification methods, such as NASBA and RT-PCR, Ovation amplification maintains representation of the starting mRNA population.

Islet Purity Matching

Our extensive experience in working with islet samples has enabled us to develop an Islet Purity Matching assay that we have found to be essential for the production of high quality microarray analysis from islets. In outline, QPCR is used to determine the relative levels of Insulin and Amylase in each sample. This then allows us to only hybridize samples that are well matched for purity in terms of the amount of exocrine contamination found in every islet preparation. In this way differential gene expression as a result of the islet preparation technique can be removed, allowing the investigator to identify to accurately determine islet gene expression profiles.