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Updated: May 26, 2026
Capsular Serotyping of Streptococcus pneumoniae Using the Quellung Reaction
Published on: February 24, 2014
Quantification of capsular polysaccharide of Streptococcus pneumoniae serotype 14 in culture broth samples
Verônica M R Gogola1, Talita S Carmo, Carolina S F Geraldo
1Laboratório de Bioprocessos, Centro de Biotecnologia, Instituto Butantan, 05503-900 São Paulo, SP, Brazil.
Insights
A new sandwich ELISA method accurately quantifies Streptococcus pneumoniae serotype 14 capsular polysaccharide (PS14). This optimized protocol identified the best PS14-producing bacterial strain for vaccine development.
Area of Science:
- Microbiology
- Immunology
- Biotechnology
Background:
- Streptococcus pneumoniae causes over a million deaths annually, primarily in developing nations.
- Pneumococcal disease prevention relies on vaccines targeting capsular polysaccharides (PS).
- Serotype 14 is a prevalent cause of pediatric pneumococcal infections globally.
Purpose of the Study:
- To develop and validate a quantification protocol for Streptococcus pneumoniae serotype 14 polysaccharide (PS14).
- To utilize the developed protocol for identifying superior PS14-producing strains of S. pneumoniae.
Main Methods:
- Evaluation of phenol-sulfuric, HPSEC, competitive ELISA, and sandwich ELISA for PS14 quantification.
- Optimization and validation of the sandwich ELISA method, determining its quantification and detection limits.
- Statistical analysis of assay performance, including intra-assay and inter-assay coefficients of variation.
Main Results:
- Sandwich ELISA demonstrated superior reproducibility, sensitivity, and resistance to interference compared to other methods.
- The optimized sandwich ELISA achieved quantification and detection limits of 0.99 ng/mL and 0.57 ng/mL, respectively.
- Strain 5287 was identified as the optimal PS14 producer, yielding 11.68 mg PS14 per biomass.
Conclusions:
- Sandwich ELISA is the preferred method for quantifying pneumococcal polysaccharides in culture broth.
- The developed protocol enables effective selection of high-yield PS14-producing S. pneumoniae strains.
- This advancement supports improved vaccine development strategies against pneumococcal disease.
Abstract:
Streptococcus pneumoniae is a major cause of mortality in underdeveloped countries, where more than one million people die from pneumococcal disease every year. Vaccines are the most efficient method for preventing the infection and are based on the capsular polysaccharide (PS) protection. The serotype 14 is the most frequent in pediatric infections worldwide. This study aimed to establish a quantification protocol for PS present in culture broth samples of S. pneumoniae serotype 14 (PS14) and use this protocol for selection of the best PS14 producer strain. Phenol-sulfuric, HPSEC, competitive ELISA, and sandwich ELISA methods were tested for PS14 quantification. Sandwich ELISA was the method with the best reproducibility and sensitivity and the least susceptible to interferences. The quantification limit and detection limit of this method were 0.99 and 0.57 ng/mL, respectively. Statistical analysis was performed to calculate the coefficient of variation (CV) intraassay (1-3% intraplate and 2-6% interplate) and interassay (11-15%) and the reproducibility in different days (CV<20%). The sandwich ELISA allows us to select, among six strains evaluated, the strain 5287 as the best PS14 producer (11.68 mg PS14/biomass) and it was shown to be the best choice for measurement of pneumococcal polysaccharides in culture broth samples.
