Detection of the Delta F508 (F508del) mutation of the cystic fibrosis geneby surface plasmon resonance and biosensor technology

In the present paper, me applied surface plasmon resonance (SPR) and biosen sor technologies for biospecific interaction analysis (BIA) to detect Delta F508 mutation (F508del) of the cystic fibrosis transmembrane regulator (CF TR) gene in both homozygous as well as heterozygous human subjects. The pro posed method is divided into three major steps. The first step is the immob ilization on a SA5 sensor chip of two biotinylated oligonucleotide probes ( one normal, N-508, and the other mutant, Delta F508) that are able to hybri dize to the CFTR gene region involved in F508del mutation, The second step consists of the molecular hybridization between the oligonucleotide probes immobilized on the sensor chips and (1) wild-type or mutant oligonucleotide s, as well as (2) single stranded DNA obtained by asymmetric polymerase cha in reaction (PCR), performed using genomic DNA from normal individuals and from F508del heterozygous and Delta 508del homozygous patients. The third, and most important, step consists of the evaluation of differential stabili ties of DNA/DNA molecular complexes generated after hybridization of normal and Delta F508 probes immobilized on the sensor chips. The results obtaine d strongly suggest that the proposed procedure employing SPR technology ena bles a one-step, nonradioactive protocol for the molecular diagnosis of F50 8del mutation of the CFTR gene. This approach could be of interest in clini cal genetics, as the hybridization step is oftenly required to detect micro deletions present within PCR products. Hum Mutat 13:390-400, 1999. (C) 1999 Wiley Liss, Inc.