title: Beta-thalassemia microelectronic chip: a fast and accurate method for mutation detection.
abstract: BACKGROUND
  beta-Thalassemia is one of the most common genetic diseases in humans. We developed an automated electronic microchip for fast and reliable detection of the nine most frequent mutations accounting for >95% of the beta-thalassemia alleles in the Mediterranean area.
  
  
  METHODS
  We developed a microchip-based assay to identify the nine most frequent mutations (cd39C>T, IVS1-110G>A, IVS1-1G>A, IVS1-6T>C, IVS2-745C>G, cd6delA, -87C>G, IVS2-1G>A, and cd8delAA) by use of the Nanogen Workstation. The biotinylated amplicon was electronically addressed on the chip to selected pads, where it remained embedded through interaction with streptavidin in the permeation layer. The DNA at each test site was then hybridized to a mixture of fluorescently labeled wild-type or mutant probes.
  
  
  RESULTS
  Assays conditions were established based on the analysis of 700 DNA samples from compound heterozygotes or homozygotes for the nine mutations. The assays were blindly validated on 250 DNA samples previously genotyped by other methods, with complete concordance of results. Alternative multiplexed formats were explored: the combination of multiplex PCR with multiple addressing and/or hybridization allowed analysis of all nine mutations in the same sample on one test site of the chip.
  
  
  CONCLUSIONS
  The open flexible platform can be designed by the user according to the local prevalence of mutations in each geographic area and can be rapidly extended to include the remaining mutations causing beta-thalassemia in other regions of the world.
authors.affiliation: Unit of Genomics for Diagnosis of Human Pathologies, Istituto di Ricovero e Cura a Carattere Scientifico Ospedale San Raffaele, Milan, Italy.
authors.forename: Barbara
authors.initials: B
authors.lastname: Foglieni
authors.forename: Laura
authors.initials: L
authors.lastname: Cremonesi
authors.forename: Maurizio
authors.initials: M
authors.lastname: Travi
authors.forename: Anna
authors.initials: A
authors.lastname: Ravani
authors.forename: Antonino
authors.initials: A
authors.lastname: Giambona
authors.forename: Maria Cristina
authors.initials: MC
authors.lastname: Rosatelli
authors.forename: Chiara
authors.initials: C
authors.lastname: Perra
authors.forename: Paolo
authors.initials: P
authors.lastname: Fortina
authors.forename: Maurizio
authors.initials: M
authors.lastname: Ferrari
mesh_terms: D005470:Fluorometry; D006801:Humans; D009154:Mutation; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction; D015203:Reproducibility of Results; D017086:beta-Thalassemia / Q000235:genetics*