DNA microarrays and flow cytometry

A DNA microarray is a small chip of glass or nylon with thousands of tiny spots of single-stranded DNA (cDNA, PCR products or short oligonucleotides) arranged in rows and columns, one spot per gene. Labelled DNA from a sample sticks (hybridises) to the spots that match it, so the activity of thousands of genes can be measured at once.

The steps:

• Extract mRNA from the cells and copy it into more stable cDNA (by reverse transcription).
• Label it with fluorescent dyes, commonly Cy3 (green) and Cy5 (red), one for each condition being compared.
• Hybridise it to the chip, then wash away anything that did not stick.
• Scan the chip and analyse the colours with software.

The software compares two conditions, for example healthy and diseased cells. Genes that are more active in one condition are up-regulated; genes that are less active are down-regulated. They show up in different colours on the scanned chip.

Flow cytometry grew from the Coulter counter (late 1950s), which counted particles by the small change in electrical resistance (impedance) each caused as it passed through a tiny hole. In a flow cytometer, cells flow in single file past one or more laser beams. Detectors in line with the beam and at right angles to it record the light scattered and the fluorescence given off by each cell, one by one.

Antibodies tagged with fluorescent dyes mark particular kinds of cells, and labelled cells, in tiny droplets, can be sorted into separate tubes by a charging electrode. Tools like these make biology more accurate, fast, quantitative and reproducible.