Note: steps taken to optimise probe specificity and signal intensity prior to field validation of the MIDTAL (Microarray for the Detection of Toxic Algae)

Abstract

A microarray for the detection of toxic algal species was developed in the European Union 7th Framework project MIDTAL. We initially tested all available fluorescence in situ hybridisation probes for toxic algae, which are normally designed to a length of 18 nt, and found that in most cases the signal was rather weak or all probes designed from the second half of the molecule were inaccessible in a microarray format because of secondary structure of the ribosomal RNA molecule We modified the length of the probes, the fragmentation of the rRNA, the stringency of the washing buffers and the length of the spacer molecules linking the probes to the glass surface of the microarray. Because of the secondary structure of the rRNA molecule, regions of the molecule can be difficult to access by the probes. Each of these modifications has improved probe accessibility and probe specificity to reduce false positives.