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Clinical Chemistry 0: clinchem.2007.098418v1, 2008; 10.1373/clinchem.2007.098418
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Received on October 11, 2007
Accepted on January 31, 2008

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. McBride 1, Francis Guy Gabriel 2, John Fordham 3, Torsten Kolind 1, Gabriela Barcenas-Morales 1, David A. Isenberg 4, Marlene Swana 1, Peter J. Delves 1, Torben Lund 1, Ian A. Cree 2, Ivan M. Roitt 1*

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

* To whom correspondence should be addressed. 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 {beta}-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 results also agreed with those obtained with conventional methods, which agreed with each other. A sample of serum from a finger stick provides a volume sufficient to perform the array assay.




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