Methods in molecular cardiology: microarray technology

It has become more and more evident that changes in expression levels of genes can play an important role in cardiovascular diseases. Specific gene expression profiles may explain, for example, the pathophysiology of myocardial hypertrophy and pump failure and may provide clues for therapeutic interventions. Knowledge of gene expression patterns can also be applied for diagnostic and prognostic purposes, in which differences in gene activity can be used for classification. DNA microarray technology has become the method of choice to simultaneously study the expression of many different genes in a single assay. Each microarray contains many thousands of different DNA sequences attached to a glass slide. The amount of messenger RNA, which is a measure of gene activity, is compared for each gene on the microarray by labelling the mRNA with different fluorescently labelled nucleotides (Cy3 or Cy5) for the test and reference samples. After hybridisation to the microarray the relative amounts of a particular gene transcript in the two samples can be determined by measuring the signal intensities for the fluorescent groups (Cy3 and Cy5) and calculating signal ratios. This paper describes the development of in-house microarray technology, using commercially available cDNA collections. Several technical approaches will be compared and an overview of the pitfalls and possibilities will be presented. The technology will be explained in the context of our project to determine gene expression differences between normal, hypertrophic and failing heart.