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Clinical and Vaccine Immunology, February 2009, p. 222-229, Vol. 16, No. 2
1071-412X/09/$08.00+0     doi:10.1128/CVI.00292-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Competitive Inhibition Flow Analysis Assay for the Non-Culture-Based Detection and Serotyping of Pneumococcal Capsular Polysaccharide {triangledown}

H. Findlow,1* G. Laher,1 P. Balmer,1 C. Broughton,2 E. D. Carrol,3 and R. Borrow1

Vaccine Evaluation Unit, Health Protection Agency North West, P.O. Box 209, Clinical Sciences Building, Manchester Royal Infirmary, Manchester, United Kingdom,1 Division of Medical Microbiology, University of Liverpool, 8th Floor, Duncan Building, Daulby Street, Liverpool, United Kingdom,2 Division of Child Health, University of Liverpool, Royal Liverpool Children's NHS Trust, Alder Hey, Eaton Road, Liverpool, United Kingdom3

Received 4 August 2008/ Returned for modification 30 September 2008/ Accepted 5 December 2008

Traditional confirmation procedures for the identification of a pneumococcal serotype require an isolate. Non-culture-based confirmation protocols are available. Some of these confirm only the presence of pneumococci, and others are capable of identifying a limited number of serotypes. The increased use of pneumococcal polysaccharide and conjugate vaccines, especially in high-risk patient groups, and the likely increase in the number of serotypes included in future versions of the conjugate vaccines have necessitated the need for improved enhanced surveillance in order to assess their impact on public health. Since 2006, a multiplexed assay has been used at the Health Protection Agency of the United Kingdom for the detection of 14 pneumococcal serotypes which requires pneumococcal serotype-specific monoclonal antibodies (MAbs). We have developed a microsphere competitive inhibition method capable of detecting 23 pneumococcal capsular polysaccharide serotypes in cerebrospinal fluid (CSF) and urine and serotyping pneumococcal suspensions, utilizing an international reference serum, 89-SF. The assay was shown to be reproducible and specific for homologous polysaccharide. Validation of the assay was performed with a selection of MAbs specific for pneumococcal capsular polysaccharide serotypes, which confirmed the specificity of the assay. Analysis of pneumolysin PCR-positive CSF samples in the competitive inhibition assay determined a serotype for 89% of the samples. The assay developed here is well suited to large-scale epidemiologic studies because the assay is simple, robust, and rapid and utilizes readily available resources.

* Corresponding author. Mailing address: Vaccine Evaluation Unit, Health Protection Agency North West, P.O. Box 209, Clinical Sciences Building, CSB2, Manchester Royal Infirmary, Oxford Road, Manchester M13 9WL, United Kingdom. Phone: 44 161 276 6791. Fax:. 44 161 276 6792. E-mail: Helen.findlow{at}hpa.org.uk

{triangledown} Published ahead of print on 17 December 2008.

Clinical and Vaccine Immunology, February 2009, p. 222-229, Vol. 16, No. 2
1071-412X/09/$08.00+0     doi:10.1128/CVI.00292-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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