title: Rapid, simple influenza RNA extraction from nasopharyngeal samples.
abstract: This report describes the development and pre-clinical testing of a new, random-access RNA sample preparation system (TruTip) for nasopharyngeal samples. The system is based on a monolithic, porous nucleic acid binding matrix embedded within an aerosol-resistant pipette tip and can be operated with single or multi-channel pipettors. Equivalent extraction efficiencies were obtained between automated QIAcube and manual TruTip methods at 10(6) gene copies influenza A per mL nasopharyngeal aspirate. Influenza A and B amended into nasopharyngeal swabs (in viral transport medium) were detected by real-time RT-PCR at approximately 745 and 370 gene copies per extraction, respectively. RNA extraction efficiency in nasopharyngeal swabs was also comparable to that obtained on an automated QIAcube instrument over a range of input concentrations; the correlation between threshold cycles (or nucleic acid recovery) for TruTip and QIAcube-purified RNA was R(2)>0.99. Preclinical testing of TruTip on blinded nasopharyngeal swab samples resulted in 98% detection accuracy relative to a clinically validated easyMAG extraction method. The physical properties of the TruTip binding matrix and ability to customize its shape and dimensions likewise make it amenable to automation and/or fluidic integration.
authors.affiliation: Akonni Biosystems, Inc., Frederick, MD 21701, USA. dchandler@akonni.com
authors.forename: Darrell P
authors.initials: DP
authors.lastname: Chandler
authors.forename: Sara B
authors.initials: SB
authors.lastname: Griesemer
authors.forename: Christopher G
authors.initials: CG
authors.lastname: Cooney
authors.forename: Rebecca
authors.initials: R
authors.lastname: Holmberg
authors.forename: Nitu
authors.initials: N
authors.lastname: Thakore
authors.forename: Becca
authors.initials: B
authors.lastname: Mokhiber
authors.forename: Phillip
authors.initials: P
authors.lastname: Belgrader
authors.forename: Christopher
authors.initials: C
authors.lastname: Knickerbocker
authors.forename: Jeanmarie
authors.initials: J
authors.lastname: Schied
authors.forename: Kirsten
authors.initials: K
authors.lastname: St George
mesh_terms: D000818:Animals; D006801:Humans; D009980:Influenza A virus / Q000235:genetics*; D009981:Influenza B virus / Q000235:genetics*; D009305:Nasopharynx / Q000821:virology*; D012367:RNA, Viral / Q000302:isolation & purification*; D013048:Specimen Handling / Q000379:methods*; D014773:Virology / Q000379:methods*
chemical_list: D012367:RNA, Viral