Simple latex agglutination method for typing pneumococci.

Lafong, A C; Crothers, E.

doi:10.1136/jcp.41.2.230

Cited by 27 publications

(29 citation statements)

References 9 publications

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“…Our assay is not only practical but also compares favorably to alternative serotyping assays currently available such as the quellung reaction (6,13,20), bacterial agglutination (2, 10), latex agglutination (9,19,21), and enzyme-linked immunosorbent assay for capsular PS (4,11). Although bacterial agglutination, latex agglutination, and enzyme-linked immunosorbent assay methods are less laborious and have an end point easier to recognize than the quellung reaction, the three assays are manual, are not multiplexed, and have a limited repertoire of identifiable serotypes.…”

Section: Discussionmentioning

confidence: 99%

Rapid Multiplex Assay for Serotyping Pneumococci with Monoclonal and Polyclonal Antibodies

Yu¹,

Lin²,

Benjamin

et al. 2005

J Clin Microbiol

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We have developed and characterized a rapid semiautomated pneumococcal serotyping system incorporating a pneumococcal lysate preparation protocol and a multiplex serotyping assay. The lysate preparation incorporates a bile solubility test to confirm pneumococcal identification that also enhances assay specificity. The multiplex serotyping assay consists of 24 assays specific for 36 serotypes: serotypes 1, 2, 3, 4, 5, 6A, 6B, 7A/7F, 8, 9L/9N, 9V, 10A/10B/39/(33C), 11A/11D/11F, 12A/12B/12F, 14, 15B/(15C), 17F, 18C, 19A, 19F, 20, 22A/22F, 23F, and 33A/33F. The multiplex assay requires a flow cytometer, two sets of latex particles coated with pneumococcal polysaccharides, and serotype-specific antibodies. Fourteen newly developed monoclonal antibodies specific for common serotypes and a pool of polyclonal rabbit sera for some of the less-common serotypes are used. The two monoclonal antibodies specific for serotypes 18C and 23F recognize serotypespecific epitopes that have not been previously described. These monoclonal antibodies make the identification of the 14 common serotypes invariant. The specificity of the serotyping assay is fully characterized with pneumococci of all known (i.e., 90) serotypes. The assay is sensitive enough to use bacterial lysates diluted 20 fold. Our serotyping system can identify not only all the serotypes in pneumococcal vaccines but also most (>90%) of clinical isolates. This system should be very useful in serotyping clinical isolates for evaluating pneumococcal vaccine efficacy.Streptococcus pneumoniae is a well-known human pathogen and a major etiologic agent for pneumonia, meningitis, otitis media, and sepsis, primarily among young children and older adults (3). S. pneumoniae is divided into 90 serotypes, based on its expression of serologically distinct carbohydrate capsules (6). Antibodies to a capsular polysaccharide (PS) provide protection against pneumococci expressing the same capsular serotype. Currently available pneumococcal vaccines contain a mixture of capsular PS of multiple serotypes. One pneumococcal vaccine (called PS vaccine) contains capsular PS from 23 commonly found serotypes (17). The most recently developed type of vaccine (called conjugate vaccine) contains capsular PS from 7 to 11 serotypes that are conjugated to a protein molecule (26). A 7-valent conjugate vaccine was introduced in 2000 for routine immunization in the United States and has reduced the incidence of invasive pneumococcal diseases in children (25).Accurate, efficient serotyping of pneumococcal isolates is important for measuring the efficacy of pneumococcal vaccines. Since the vaccine-induced antibodies provide serotypespecific protection, following the introduction of a new pneumococcal vaccine, pneumococci expressing the serotypes included in the vaccine would become less common while the prevalence of the pneumococci expressing nonvaccine serotypes would stay the same. As a result, a reduction in vaccine serotypes reflects the vaccine efficacy. In some cases, pneumococci expressing the non...

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Section: Discussionmentioning

confidence: 99%

Rapid Multiplex Assay for Serotyping Pneumococci with Monoclonal and Polyclonal Antibodies

Yu¹,

Lin²,

Benjamin

et al. 2005

J Clin Microbiol

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“…Commercial pneumococcal latex agglutination reagents are available, but they currently do not differentiate all known pneumococcal serotypes A positive latex reaction occurs when specific antibodies bound to the latex particles attach to antigens on the polysaccharide capsule of the pneumococcal isolate 18,19 . The latex particles attached to the antibodies allow the specific antibody-antigen reaction to be visualized without magnification.…”

Section: Discussionmentioning

confidence: 99%

“…In the method described here, the antibody is attached to latex particles via passive adsorption 15,19 , so a critical variable is the concentration of antibody used, which determines how well the surface of the latex particles is covered and the subsequent alignment of the antibody active sites 15,20 . Consequently, if insufficient or excess antibody is attached to the latex particles, the antibody-antigen reactions will not be optimal, and unsatisfactory reagents may be produced 15,20,21 .…”

Section: Discussionmentioning

confidence: 99%

Capsular Serotyping of <em>Streptococcus pneumoniae</em> by Latex Agglutination

Porter

Ortika

Satzke

2014

JoVE

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Latex agglutination reagents are widely used in microbial diagnosis, identification and serotyping. Streptococcus pneumoniae (the pneumococcus) is a major cause of morbidity and mortality world-wide. Current vaccines target the pneumococcal capsule, and there are over 90 capsular serotypes. Serotyping pneumococcal isolates is therefore important for assessing the impact of vaccination programs and for epidemiological purposes. The World Health Organization has recommended latex agglutination as an alternative method to the 'gold standard' Quellung test for serotyping pneumococci. Latex agglutination is a relatively simple, quick and inexpensive method; and is therefore suitable for resource-poor settings as well as laboratories with high-volume workloads. Latex agglutination reagents can be prepared in-house utilizing commercially-sourced antibodies that are passively attached to latex particles. This manuscript describes a method of production and quality control of latex agglutination reagents, and details a sequential testing approach which is time-and cost-effective.This method of production and quality control may also be suitable for other testing purposes.

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“…Dot blot assay. To investigate discrepant results, this assay was performed as described previously (5) using pneumococcal antisera from Statens Serum Institut for the following serogroups or serotypes: 1,4,5,6,7,8,9,11,12,14,18, and 23. Monoclonal antibodies specific for 6A (Hyp6AM3) and 18C (Hyp18CM1) serotypes were also used in some cases.…”

Section: Methodsmentioning

confidence: 99%

Validation of a Multiplex Pneumococcal Serotyping Assay with Clinical Samples

et al. 2006

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) were noted, but 46 were due to nonreproducible technical problems or insufficient growth of the pneumococci. All of the isolates grew well for a second test, and therefore, the culture medium used for the multibead assay is adequate. The discrepancies persisted for eight isolates, involving the 6A, 11A, and 18C serotypes. Additional studies of the eight isolates showed that the discrepancies were due to differences in the reagents used in the multibead or Neufeld tests for these three serotypes. For instance, five isolates were typed as 6A with the Neufeld test but as nontypeable by the multibead assay. Selection of another new monoclonal antibody (Hyp6AG1) for the multibead assay resulted in all five discrepant isolates typing as 6A. This finding indicates the validity of the multibead assay and emphasizes the need to validate any new pneumococcal serotyping assay with a large number of clinical isolates from different locations. It also suggests the presence of serological subtypes among isolates expressing the 6A serotype.

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Simple latex agglutination method for typing pneumococci.

Cited by 27 publications

References 9 publications

Rapid Multiplex Assay for Serotyping Pneumococci with Monoclonal and Polyclonal Antibodies

Rapid Multiplex Assay for Serotyping Pneumococci with Monoclonal and Polyclonal Antibodies

Capsular Serotyping of <em>Streptococcus pneumoniae</em> by Latex Agglutination

Validation of a Multiplex Pneumococcal Serotyping Assay with Clinical Samples

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