title: Development of an automated DNA purification module using a micro-fabricated pillar chip.
abstract: We present a fully automated DNA purification module comprised of a micro-fabricated chip and sequential injection analysis system that is designed for use within autonomous instruments that continuously monitor the environment for the presence of biological threat agents. The chip has an elliptical flow channel containing a bed (3.5 x 3.5 mm) of silica-coated pillars with height, width and center-to-center spacing of 200, 15, and 30 microm, respectively, which provides a relatively large surface area (ca. 3 cm(2)) for DNA capture in the presence of chaotropic agents. We have characterized the effect of various fluidic parameters on extraction performance, including sample input volume, capture flow rate, and elution volume. The flow-through design made the pillar chip completely reusable; carryover was eliminated by flushing lines with sodium hypochlorite and deionized water between assays. A mass balance was conducted to determine the fate of input DNA not recovered in the eluent. The device was capable of purifying and recovering Bacillus anthracis genomic DNA (input masses from 0.32 to 320 pg) from spiked environmental aerosol samples, for subsequent analysis using polymerase chain reaction-based assays.
authors.affiliation: Lawrence Livermore National Laboratory, Livermore, CA 94550, USA.
authors.forename: Benjamin J
authors.initials: BJ
authors.lastname: Hindson
authors.forename: Dora M
authors.initials: DM
authors.lastname: Gutierrez
authors.forename: Kevin D
authors.initials: KD
authors.lastname: Ness
authors.forename: Anthony J
authors.initials: AJ
authors.lastname: Makarewicz
authors.forename: Thomas R
authors.initials: TR
authors.lastname: Metz
authors.forename: Ujwal S
authors.initials: US
authors.lastname: Setlur
authors.forename: William B
authors.initials: WB
authors.lastname: Benett
authors.forename: Jeffrey M
authors.initials: JM
authors.lastname: Loge
authors.forename: Bill W
authors.initials: BW
authors.lastname: Colston
authors.forename: Paul S
authors.initials: PS
authors.lastname: Francis
authors.forename: Neil W
authors.initials: NW
authors.lastname: Barnett
authors.forename: John M
authors.initials: JM
authors.lastname: Dzenitis
mesh_terms: D000336:Aerosols; D000393:Air Pollutants / Q000032:analysis*; D000818:Animals; D001408:Bacillus anthracis / Q000235:genetics*; D004269:DNA, Bacterial / Q000032:analysis*; D004784:Environmental Monitoring / Q000379:methods*; D004867:Equipment Design; D017022:Flow Injection Analysis; D006801:Humans; D008832:Microchemistry; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction / Q000379:methods
chemical_list: D000336:Aerosols; D000393:Air Pollutants; D004269:DNA, Bacterial