Rapid, simple influenza RNA extraction from nasopharyngeal samples

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...

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Bibliographic Details
Published in:Journal of virological methods 2012-07, Vol.183 (1), p.8-13
Main Authors: Chandler, Darrell P., Griesemer, Sara B., Cooney, Christopher G., Holmberg, Rebecca, Thakore, Nitu, Mokhiber, Becca, Belgrader, Phillip, Knickerbocker, Christopher, Schied, Jeanmarie, St. George, Kirsten
Format: Article
Language:English
Subjects:
Animals
Automation
Biological and medical sciences
Fundamental and applied biological sciences. Psychology
genes
Humans
influenza
Influenza A
Influenza A virus - genetics
Influenza B virus - genetics
Integration
Microbiology
Nasopharyngeal aspirate
Nasopharyngeal swab
Nasopharynx - virology
nucleic acids
physical properties
Polymerase chain reaction
reverse transcriptase polymerase chain reaction
RNA
RNA, Viral - isolation & purification
Sample preparation
Specimen Handling - methods
Techniques used in virology
Virology
Virology - methods
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Summary: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 106 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 R2>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.
ISSN:0166-0934
1879-0984
DOI:10.1016/j.jviromet.2012.03.002