title: Integrated amplification microarray system in a lateral flow cell for warfarin genotyping from saliva.
abstract: BACKGROUND
  Genetic polymorphisms in the CYP2C9 and VKORC1 genes have been linked to sensitivity of the anticoagulant drug warfarin. The aim of this study is to demonstrate a method for warfarin sensitivity genotyping using gel element microarray technology in a simplified workflow from sample collection to analysis and detection.
  
  
  METHODS
  We developed an integrated amplification microarray system combining PCR amplification, target labeling, and microarray hybridization within a single, closed-amplicon "lateral flow cell" for genotyping three single nucleotide polymorphisms (SNPs) that influence warfarin response. We combined nucleic acid extraction of saliva using the TruTip technology together with the lateral flow cell assay and with fully automated array detection and analysis.
  
  
  RESULTS
  The analytical performance of the assay was tested using 20 genotyped human genomic DNA samples and found to be sensitive down to 330 input genomic copies (1 ng). A follow-up pre-clinical evaluation was performed with 65 blinded saliva samples and the genotyping results were in agreement with those determined by bidirectional sequencing.
  
  
  CONCLUSIONS
  Combined with the use of non-invasive saliva samples, rapid DNA extraction, the lateral flow cell, automatic imaging and data analysis provides a simple and fast sample-to-answer microarray test for warfarin sensitivity genotyping.
authors.affiliation: Akonni Biosystems, Inc., 400 Sagner Avenue, Suite 300, Frederick, MD 21701, United States. Electronic address: tsebastian@akonni.com.
authors.forename: Thomas
authors.initials: T
authors.lastname: Sebastian
authors.affiliation: Akonni Biosystems, Inc., 400 Sagner Avenue, Suite 300, Frederick, MD 21701, United States.
authors.forename: Christopher G
authors.initials: CG
authors.lastname: Cooney
authors.affiliation: Akonni Biosystems, Inc., 400 Sagner Avenue, Suite 300, Frederick, MD 21701, United States.
authors.forename: Jennifer
authors.initials: J
authors.lastname: Parker
authors.affiliation: Akonni Biosystems, Inc., 400 Sagner Avenue, Suite 300, Frederick, MD 21701, United States.
authors.forename: Peter
authors.initials: P
authors.lastname: Qu
authors.affiliation: Akonni Biosystems, Inc., 400 Sagner Avenue, Suite 300, Frederick, MD 21701, United States.
authors.forename: Alexander
authors.initials: A
authors.lastname: Perov
authors.affiliation: Akonni Biosystems, Inc., 400 Sagner Avenue, Suite 300, Frederick, MD 21701, United States.
authors.forename: Julia B
authors.initials: JB
authors.lastname: Golova
authors.affiliation: DNA Genotek, Inc., 2 Beaverbrook Road, Kanata, Ontario K2K 1L1, Canada.
authors.forename: Lindsay
authors.initials: L
authors.lastname: Pozza
authors.affiliation: DNA Genotek, Inc., 2 Beaverbrook Road, Kanata, Ontario K2K 1L1, Canada.
authors.forename: Rafal M
authors.initials: RM
authors.lastname: Iwasiow
authors.affiliation: Akonni Biosystems, Inc., 400 Sagner Avenue, Suite 300, Frederick, MD 21701, United States.
authors.forename: Rebecca
authors.initials: R
authors.lastname: Holmberg
mesh_terms: D000483:Alleles; D001189:Aryl Hydrocarbon Hydroxylases / Q000235:genetics; D065729:Cytochrome P-450 CYP2C9; D060005:Genotyping Techniques / Q000379:methods*; D006801:Humans; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods*; D020641:Polymorphism, Single Nucleotide; D012463:Saliva / Q000166:cytology* / Q000378:metabolism*; D018511:Systems Integration; D064417:Vitamin K Epoxide Reductases / Q000235:genetics; D014859:Warfarin / Q000494:pharmacology*
chemical_list: D014859:Warfarin; C450260:CYP2C9 protein, human; D065729:Cytochrome P-450 CYP2C9; D001189:Aryl Hydrocarbon Hydroxylases; C578523:VKORC1 protein, human; D064417:Vitamin K Epoxide Reductases
keywords: CYP2C9; Lateral flow cell; Microarray; PCR; SNP; Saliva; VKORC1; Warfarin; cytochrome P-450 variant 2C9; polymerase chain reaction; single-nucleotide polymorphism; vitamin K epoxide reductase complex-1