Clinical Chemistry
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Clinical Chemistry 54: 883-890, 2008. First published March 20, 2008; 10.1373/clinchem.2007.098418
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(Clinical Chemistry. 2008;54:883-890.)
© 2008 American Association for Clinical Chemistry, Inc.

Clinical Immunology

Screening Autoantibody Profiles in Systemic Rheumatic Disease with a Diagnostic Protein Microarray That Uses a Filtration-Assisted Nanodot Array Luminometric Immunoassay (NALIA)

Jeffrey D. McBride1, Francis Guy Gabriel2, John Fordham3, Torsten Kolind1, Gabriela Barcenas-Morales1, David A. Isenberg4, Marlene Swana1, Peter J. Delves1, Torben Lund1, Ian A. Cree2 and Ivan M. Roitt1,a

1 Department of Immunology and Molecular Pathology, University College London, London, UK; 2 Translational Oncology Research Centre, University of Portsmouth and Department of Histopathology, Queen Alexandra Hospital, Portsmouth, UK; 3 Department of Physics and Astronomy, University College London, London, UK; 4 Department of Medicine, University College London, London, UK.

aAddress correspondence to this author at: Department of Immunology and Molecular Pathology, UCL, 46 Cleveland St., London W1T 4JF, UK. Fax 44-20-7679-9400; e-mail i.roitt{at}ucl.ac.uk.

Background: We developed a cost-efficient modular system for multiplex analysis of the multiple autoantibodies that characterize systemic rheumatoid diseases.

Methods: The nanodot array luminometric immunoassay (NALIA) system consists of conventional 96-well membrane-bottomed plates in which antigens or antibodies are adsorbed onto the underside of the membrane. Current arrays use a 5 x 5 format (25 dots/well), which allows 10 analytes to be measured in duplicate: double-stranded DNA (dsDNA), centromere protein B (CENP-B), PCNA, Sm, Sm ribonucleoprotein (Sm-RNP), U1-snRNP, Scl70, SSA/Ro, SSB/La, Jo-1, and controls. The test fluid, control sera, and subsequent reagents are drawn through the membrane. The captured analytes are quantified by monitoring chemiluminescence with a charge-coupled device (CCD) and analyzed with commercial array software.

Results: The assay can detect <20 x 103 IU/L of anti-dsDNA. The interwell CV was 10%–14%. There was an 83% concordance ({kappa} = 0.56) between the NALIA results obtained for anti-dsDNA assayed by β-testing in a routine immunology diagnostic laboratory and the results obtained with a conventional ELISA reagent set. The concordance values for Ro, La, Sm, and RNP were 98% ({kappa}, 0.92), 93% ({kappa}, 0.41), 97% ({kappa}, 0.62), and 97% ({kappa}, 0.73), respectively.

Conclusion: The NALIA approach promises to provide a highly economical platform for a wide range of applications that require assays of multiple analytes. The degree of concordance of our results with a conventional reagent set was no less than that occurring between different commercial products. A sample of serum from a finger stick provides a volume sufficient to perform the array assay.






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