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Journal of Clinical Microbiology, August 2006, p. 2863-2871, Vol. 44, No. 8
0095-1137/06/$08.00+0     doi:10.1128/JCM.00134-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Experimental Evaluation of the FluChip Diagnostic Microarray for Influenza Virus Surveillance

Department of Chemistry and Biochemistry, The University of Colorado at Boulder, UCB #215, Boulder, CO 80309,¹ Influenza Branch, Centers for Disease Control and Prevention, 1600 Clifton Rd., Atlanta, GA 30333,² InDevR, LLC, 2100 Central Ave., Suite 106, Boulder, CO 80301³

Received 20 January 2006/ Returned for modification 10 April 2006/ Accepted 14 May 2006

Global surveillance of influenza is critical for improvements in disease management and is especially important for early detection, rapid intervention, and a possible reduction of the impact of an influenza pandemic. Enhanced surveillance requires rapid, robust, and inexpensive analytical techniques capable of providing a detailed analysis of influenza virus strains. Low-density oligonucleotide microarrays with highly multiplexed "signatures" for influenza viruses offer many of the desired characteristics. However, the high mutability of the influenza virus represents a design challenge. In order for an influenza virus microarray to be of utility, it must provide information for a wide range of viral strains and lineages. The design and characterization of an influenza microarray, the FluChip-55 microarray, for the relatively rapid identification of influenza A virus subtypes H1N1, H3N2, and H5N1 are described here. In this work, a small set of sequences was carefully selected to exhibit broad coverage for the influenza A and B viruses currently circulating in the human population as well as the avian A/H5N1 virus that has become enzootic in poultry in Southeast Asia and that has recently spread to Europe. A complete assay involving extraction and amplification of the viral RNA was developed and tested. In a blind study of 72 influenza virus isolates, RNA from a wide range of influenza A and B viruses was amplified, hybridized, labeled with a fluorophore, and imaged. The entire analysis time was less than 12 h. The combined results for two assays provided the absolutely correct types and subtypes for an average of 72% of the isolates, the correct type and partially correct subtype information for 13% of the isolates, the correct type only for 10% of the isolates, false-negative signals for 4% of the isolates, and false-positive signals for 1% of the isolates. In the overwhelming majority of cases in which incomplete subtyping was observed, the failure was due to the nucleic acid amplification step rather than limitations in the microarray.

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