CVI Accepts, published online ahead of print on 28 October 2009

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Clin. Vaccine Immunol. doi:10.1128/CVI.00248-09
Copyright (c) 2009, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Development and Analytical Validation of Immunoassay for Quantifying Serum Anti-Pertussis Toxin Antibodies Resulting from Bordetella pertussis Infection

Sandra L. Menzies^*, Vijay Kadwad, Lucia Pawloski, Tsai-Lien Lin, Andrew L. Baughman, Monte Martin, Maria Lucia C. Tondella, Bruce D. Meade, and the Pertussis Assay Working Group

Division of Bacterial, Parasitic, and Allergenic Products, Center for Biologics Evaluation and Research, FDA, Rockville, Maryland; Division of Bacterial Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia; Division of Biostatistics, Center for Biologics Evaluation and Research, FDA, Rockville, Maryland; Immunoassay Department, BRIT, India; Meade Biologics, LLC, Hillsborough, NC

^* To whom correspondence should be addressed. Email: Sandra.Menzies{at}fda.hhs.gov.

Adequately sensitive and specific methods to diagnose pertussis in adolescents and adults are not widely available. Currently, no Food and Drug Administration approved diagnostic assays are available for the serodiagnosis of Bordetella pertussis. Since concentrations of B. pertussis-specific antibodies tend to be high during the later-phases of disease, a simple, rapid, easily transferable serodiagnostic test was developed. This paper describes test development, initial evaluation of a prototype kit enzyme-linked immunosorbent assay (ELISA) in an inter-laboratory collaborative study, and the analytical validation. The data presented here demonstrate that the kit met all pre-specified criteria for precision, linearity, and accuracy for samples with anti-pertussis toxin (PT) immunoglobulin G (IgG) antibody concentrations in the range of 50 to 150 ELISA units (EU)/mL, the range believed to be most relevant for serodiagnosis. The assay met the precision and linearity criteria for a wider range, namely from 50 to 200 EU/mL, however, the accuracy criterion was not met at 200 EU/mL. When the newly adopted World Health Organization International Standard for pertussis antiserum (human) reference reagent was used to evaluate accuracy, the accuracy criteria were met from 50 to 200 international units (IU)/mL. In conclusion, the IgG anti-PT ELISA assay met all assay validation parameters within the range considered most relevant for serodiagnosis. This ELISA was developed and analytically validated to be a user-friendly kit that can be used in both qualitative and quantitative formats. The technology for producing the kit is transferrable to public health laboratories.