Title: The application of IgM in an xMAP suspension assay using recombinant Alt a1 as a model target protein

Authors: Makonnen Abebe, Kathy Nguyen, Azam Tayabali, Wendy Decker, Veena Kumar, Serdal Sevinc, Hari Vijay

Addresses: Environmental Health Science Research Bureau, Health Canada, Tunney's Pasture, Ottawa, K1A 0K9, ON, Canada. ' Environmental Health Science Research Bureau, Health Canada, Tunney's Pasture, Ottawa, K1A 0K9, ON, Canada. ' Environmental Health Science Research Bureau, Health Canada, Tunney's Pasture, Ottawa, K1A 0K9, ON, Canada. ' Public Health Agency of Canada, National HIV and Retrovirology Laboratory, Health Canada, Tunney's Pasture, Ottawa, K1A 0K9, ON, Canada. ' Environmental Health Science Research Bureau, Health Canada, Tunney's Pasture, Ottawa, K1A 0K9, ON, Canada.' Environmental Health Science Research Bureau, Health Canada, Tunney's Pasture, Ottawa, K1A 0K9, ON, Canada. ' Environmental Health Science Research Bureau, Health Canada, Tunney's Pasture, Ottawa, K1A 0K9, ON, Canada

Abstract: The potential of Immunoglobulin M (IgM) in an Multi Analyte Profiling (xMAP) multiplex suspension assay was determined using Alt a1, a major allergen of Alternaria alternata, as the model target protein. LumAvidin (LAM) and carboxylated (CAM) microspheres, from LUMINEX™ Corporation, were compared. The reproducible standard curves for concentrations between 1 pg/mL and 10 ng/mL and 3 pg/mL and 300 ng/mL of the allergen were generated by the LAM assay, allowing for the quantitation of an unknown test sample. The success of the LAM assay is dependent on avidin molecules, initially bound on the microsphere, along with the use of a phycoerythrin-coupled secondary IgM. The standard curves generated by the CAM assay for the two allergen concentrations were erratic, falling short of quantifying the test sample. This is the first time a monoclonal IgM is successfully applied as a primary and secondary antibody in the xMAP format. The routine application of this assay will be consolidated as new IgM-based xMAP diagnostics become available.

Keywords: immunoglobulin M; monoclonal IgM; multi analyte profiling; xMAP diagnostics; LumAvidin microspheres; carboxylated microspheres; Alt a1; LAM assay; CAM assay; multiplex suspension assay; primary antibodies; secondary antibodies; allergens; model target protein.

DOI: 10.1504/IJIS.2009.023558

International Journal of Immunological Studies, 2009 Vol.1 No.1, pp.1 - 12

Published online: 28 Feb 2009 *

Full-text access for editors Full-text access for subscribers Purchase this article Comment on this article