This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Marchese, R. D.
Right arrow Articles by Sikkema, D.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Marchese, R. D.
Right arrow Articles by Sikkema, D.

 Previous Article  |  Next Article 

Clinical and Vaccine Immunology, March 2009, p. 387-396, Vol. 16, No. 3
1071-412X/09/$08.00+0     doi:10.1128/CVI.00415-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Optimization and Validation of a Multiplex, Electrochemiluminescence-Based Detection Assay for the Quantitation of Immunoglobulin G Serotype-Specific Antipneumococcal Antibodies in Human Serum{triangledown}

Rocio D. Marchese,1* Derek Puchalski,1 Pamela Miller,2 Joseph Antonello,2 Olivia Hammond,1 Tina Green,2 Leonard J. Rubinstein,3 Michael J. Caulfield,1 and Daniel Sikkema1,{dagger}

Wayne Clinical Support, Merck Research Laboratories, Wayne, Pennsylvania,1 Non-Clinical Statistics, Merck Research Laboratories, West Point, Pennsylvania,2 ID/Vaccines, Merck Research Laboratories, Upper Gwynedd, Pennsylvania3

Received 11 November 2008/ Returned for modification 3 December 2008/ Accepted 12 January 2009

Pneumovax 23 consists of a mixture of highly purified capsular polysaccharides (Ps) from 23 of the most prevalent serotypes of Streptococcus pneumoniae. Testing of vaccine immunogenicity has been historically performed on the enzyme-linked immunosorbent assay (ELISA) platform, validated to measure immunoglobulin G (IgG) antibodies to all 23 serotypes included in Pneumovax 23. In order to significantly improve the throughput of this form of testing, we have developed and validated a direct binding electrochemiluminescence (ECL)-based multiplex assay that can measure the antibody response in human serum to eight serotypes within a single microtiter well. The pneumococcal (Pn) ECL assay is based on the Meso Scale Discovery (MSD) technology which utilizes a Sulfo-Tag-labeled anti-human IgG antibody that emits light upon electrochemical stimulation. The Pn ECL assay exhibits a wide dynamic range and provides the ability to read concentrations down to the minimum reported concentration in the Merck ELISA (0.1 µg/ml). Cross-reactivity assessment using type-specific monoclonal antibodies showed no cross talk between antigen spots within a well. By use of the WHO Pn sample reference panel, the results obtained with the Pn ECL assay were compared to the results obtained with the international Pn ELISA. The results for the Pn ECL assay satisfied the WHO-recommended acceptance criterion for concordance for all seven serotypes with published Pn ELISA values, and the overall correlation (r value) across the seven serotypes was 0.994. The MSD methodology has great potential to be extremely useful for simultaneously quantitating IgG responses to several Pn serotypes while conserving serum volumes and laboratory testing time.

* Corresponding author. Mailing address: ID/Vaccines Division, Merck & Co. Inc., P.O. Box 1000, UG3CD28, North Wales, PA 19454-1099. Phone: (267) 305-8492. Fax: (267) 305-6515. E-mail: rocio_marchese{at}merck.com

{triangledown} Published ahead of print on 21 January 2009.

{dagger} Present address: Sanofi Pasteur, Mailstop B55-1102, Discovery Drive, Swiftwater, PA 18370.

Clinical and Vaccine Immunology, March 2009, p. 387-396, Vol. 16, No. 3
1071-412X/09/$08.00+0     doi:10.1128/CVI.00415-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»