Peptide nucleic acids and biosensor technology for real-time detection of the cystic fibrosis W1282X mutation by surface plasmon resonance

in this paper we demonstrate that peptide nucleic acids (PNAs) are excellen t probes able to detect the W1282X point mutation of the cystic fibrosis (C F) gene when biospecific interaction analysis (BIA) by surface plasmon reso nance (SPR) and biosensor technologies is performed. The results reported h ere suggest that BIA is an easy, fast, and automatable approach for detecti ng mutations of OF, allowing real-time monitoring of hybridization between 9-mer OF PNA probes and target biotinylated POR products generated from hea lthy, heterozygous subjects and homozygous W1282X samples and immobilized o n streptavidin-coated sensor chips. This method is, to our knowledge, the f irst application of PNAs, BIA, and SPR to a human hereditary mutation, and demonstrates the feasibility of these approaches for discriminating between normal and mutated target DNA. We like to point out that the procedure des cribed in this paper is rapid and informative; results are obtained within a few minutes. This could be of great interest for molecular pre-implantati on diagnosis to discriminate homozygous OF embryos from heterozygous and he althy embryos. Other advantages of the methodology described in the present paper are (a) that it is a nonradioactive methodology and (b) that gel ele ctrophoresis and/or dot-spot analysis are not required. More importantly, t he demonstration that SPR-based BIA could be associated with microarray tec hnology allows us to hypothesize that the method described in the present p aper could be used for the development of a protocol employing multispottin g on SPR biosensors of many CF-PCR products and a real-time simultaneous an alysis of hybridization to PNA probes. These results are in line with the c oncept that SPR could be an integral part of a fully automated diagnostic s ystem based on the use of laboratory workstations, biosensors, and arrayed biosensors for DNA isolation, preparation of PCR reactions, and identificat ion of point mutations.