scholarly journals Evaluation of the Specificity of Pneumococcal Polysaccharide Enzyme-Linked Immunosorbent Assay and the Effect of Serum Adsorption Based on Standard Pneumococcal Serogroup- or Serotype-Specific Rabbit Antisera

2009 ◽  
Vol 16 (9) ◽  
pp. 1279-1284 ◽  
Author(s):  
Hans-Christian Slotved ◽  
Christina Guttmann ◽  
Charlotte Demuth Pedersen ◽  
Jasper Neergaard Jacobsen ◽  
Karen Angeliki Krogfelt
Keyword(s):  
Cell Wall ◽  
Streptococcus Pneumoniae ◽  
Human Serum ◽  
Immunosorbent Assay ◽  
Capsular Polysaccharide ◽  
Enzyme Linked Immunosorbent Assay ◽  
Capsular Polysaccharides ◽  
Who Guidelines ◽  
Specific Antibodies ◽  
Specific Rabbit

ABSTRACT Worldwide, Streptococcus pneumoniae (pneumococcus) is a major cause of morbidity and mortality, especially in infants and elderly people. Pneumococcal capsular polysaccharides are well characterized, and more than 90 different serotypes have been identified. Serotype-specific antibodies against the capsular polysaccharide are produced during infection. Detection of antibodies against pneumococci by enzyme-linked immunosorbent assay (ELISA) is performed according to WHO guidelines, using antigens provided by ATCC. However, testing the ELISA for specificity is challenging due to the difficulty in obtaining human naïve serum with pneumococcal antibodies as well as human serum with antibodies against a single serotype. The application of well-defined serotype-specific sera produced in animals to evaluate the specificity of the ATCC antigens and the effect of adsorption with cell wall and 22F polysaccharides has not been performed before, to our knowledge. In this study, the specificity of ATCC antigens (serotypes 4, 6B, 9V, 14, 18C, 19F, and 23F) was tested by using commercial serotype-, serogroup-, and pool-specific pneumococcal rabbit antisera.

scholarly journals Assignment of Immunoglobulin G1 and G2 Concentrations to Pneumococcal Capsular Polysaccharides 3, 6B, 14, 19F, and 23F in Pneumococcal Reference Serum 89-SF

1998 ◽  
Vol 5 (4) ◽  
pp. 561-566 ◽  
Author(s):  
Anu Soininen ◽  
Ilkka Seppälä ◽  
Tomi Wuorimaa ◽  
Helena Käyhty
Keyword(s):  
Immunosorbent Assay ◽  
Capsular Polysaccharide ◽  
Enzyme Linked Immunosorbent Assay ◽  
Capsular Polysaccharides ◽  
Serum Pool ◽  
Reference Serum

ABSTRACT We describe standardization of an enzyme-linked immunosorbent assay (ELISA) for measuring immunoglobulin G1 (IgG1) and IgG2 concentrations of antibodies to pneumococcal capsular polysaccharide (Pnc PS). The ELISA uses a human pneumococcal reference serum pool, lot 89-SF, as a reference. IgG1 and IgG2 concentrations were assigned to antibodies to Pnc PS serotypes 3, 6B, 14, 19F, and 23F in 89-SF by ELISA using affinity-purified monoisotypic IgG1 and IgG2 preparations. The sum of IgG1 and IgG2 concentrations in 89-SF agrees well with the previously assigned IgG concentrations. The IgG1 and IgG2 values in 89-SF were used to measure antibodies to Pnc PS 6B, 14, and 23F in adult pre- and postimmunization sera and the sum of IgG1 and IgG2 concentrations correlated well with the IgG values. Furthermore, the IgG2/IgG1 ratio did not affect the detection of IgG1, the isotype usually represented by a lower concentration.

scholarly journals Use of an Inhibition Enzyme-Linked Immunosorbent Assay for Quantification of Capsular Polysaccharide or Proteins in Vaccines

2007 ◽  
Vol 14 (3) ◽  
pp. 323-327 ◽  
Author(s):  
Thomas J. Inzana ◽  
Anna Champion
Keyword(s):  
Immunosorbent Assay ◽  
Capsular Polysaccharide ◽  
Actinobacillus Pleuropneumoniae ◽  
Enzyme Linked Immunosorbent Assay ◽  
Specific Antibodies ◽  
Whole Cells

ABSTRACT An inhibition enzyme-linked immunosorbent assay (ELISA) is described for quantification of capsular polysaccharide or proteins in vaccines and other samples containing whole cells or extracts of Actinobacillus pleuropneumoniae. The assay can be used to quantify any antigen that can be purified and for which highly specific antibodies are not available. The assay can be carried out by any laboratory capable of performing an ELISA.

scholarly journals Synthetic 6B Di-, Tri-, and Tetrasaccharide-Protein Conjugates Contain Pneumococcal Type 6A and 6B Common and 6B- Specific Epitopes That Elicit Protective Antibodies in Mice

2001 ◽  
Vol 69 (2) ◽  
pp. 787-793 ◽  
Author(s):  
Wouter T. M. Jansen ◽  
Sietske Hogenboom ◽  
Mark J. L. Thijssen ◽  
Johannis P. Kamerling ◽  
Johannes F. G. Vliegenthart ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Immunosorbent Assay ◽  
Capsular Polysaccharide ◽  
Keyhole Limpet Hemocyanin ◽  
Enzyme Linked Immunosorbent Assay ◽  
Protective Capacity ◽  
Protein Conjugates ◽  
Protective Antibodies

ABSTRACT The immunogenicity and protective capacity of Streptococcus pneumoniae 6B capsular polysaccharide (PS)-derived synthetic phosphate-containing disaccharide (Rha-ribitol-P-), trisaccharide (ribitol-P-Gal-Glc-), and tetrasaccharide (Rha-ribitol-P-Gal-Glc-)-protein conjugates in rabbits and mice were studied. In rabbits, all saccharides conjugated to keyhole limpet hemocyanin (KLH) evoked high levels of pneumococcal (Pn) type 6B antibodies that facilitated type-specific phagocytosis. Unlike the disaccharide rabbit antisera, tri- and tetrasaccharide rabbit antisera also reacted with 6A PS in an enzyme-linked immunosorbent assay (ELISA) and promoted phagocytosis of 6A pneumococci. All these rabbit antisera passively protected mice against a Pn 6B challenge. The disaccharide conjugate-induced antiserum, however, failed to protect mice against a 6A challenge. In mice, phagocytic and protective anti-Pn 6B antibodies were only induced by the tetrasaccharide conjugate and not by PS 6B or PS 6B-protein conjugates. These antibodies did not cross-react with 6A PS in ELISA and were unable to phagocytize 6A pneumococci. In conclusion, the disaccharide and tetrasaccharide conjugates already contain epitopes capable of inducing 6B-specific, fully protective antibodies in rabbits and mice, respectively.

scholarly journals Antibodies to Streptococcus pneumoniae Capsular Polysaccharide Enhance Pneumococcal Quorum Sensing

mBio ◽  
2011 ◽  
Vol 2 (5) ◽  
Author(s):  
Masahide Yano ◽  
Shruti Gohil ◽  
J. Robert Coleman ◽  
Catherine Manix ◽  
Liise-anne Pirofski
Keyword(s):  
Monoclonal Antibodies ◽  
Streptococcus Pneumoniae ◽  
Quorum Sensing ◽  
Direct Effect ◽  
Pneumococcal Disease ◽  
Effector Cell ◽  
Capsular Polysaccharide ◽  
Genetic Exchange ◽  
Content Type ◽  
Specific Antibodies

ABSTRACTThe use of pneumococcal capsular polysaccharide (PPS)-based vaccines has resulted in a substantial reduction in invasive pneumococcal disease. However, much remains to be learned about vaccine-mediated immunity, as seven-valent PPS-protein conjugate vaccine use in children has been associated with nonvaccine serotype replacement and 23-valent vaccine use in adults has not prevented pneumococcal pneumonia. In this report, we demonstrate that certain PPS-specific monoclonal antibodies (MAbs) enhance the transformation frequency of two differentStreptococcus pneumoniaeserotypes. This phenomenon was mediated by PPS-specific MAbs that agglutinate but do not promote opsonic effector cell killing of the homologous serotypeinvitro. Compared to the autoinducer, competence-stimulating peptide (CSP) alone, transcriptional profiling of pneumococcal gene expression after incubation with CSP and one such MAb to the PPS of serotype 3 revealed changes in the expression of competence (com)-related and bacteriocin-like peptide (blp) genes involved in pneumococcal quorum sensing. This MAb was also found to induce a nearly 2-fold increase in CSP2-mediated bacterial killing or fratricide. These observations reveal a novel, direct effect of PPS-binding MAbs on pneumococcal biology that has important implications for antibody immunity to pneumococcus in the pneumococcal vaccine era. Taken together, our data suggest heretofore unsuspected mechanisms by which PPS-specific antibodies could affect genetic exchange and bacterial viability in the absence of host cells.IMPORTANCECurrent thought holds that pneumococcal capsular polysaccharide (PPS)-binding antibodies protect against pneumococcus by inducing effector cell opsonic killing of the homologous serotype. While such antibodies are an important part of how pneumococcal vaccines protect against pneumococcal disease, PPS-specific antibodies that do not exhibit this activity but are highly protective against pneumococcus in mice have been identified. This article examines the effect of nonopsonic PPS-specific monoclonal antibodies (MAbs) on the biology ofStreptococcus pneumoniae. The results showed that in the presence of a competence-stimulating peptide (CSP), such MAbs increase the frequency of pneumococcal transformation. Further studies with one such MAb showed that it altered the expression of genes involved in quorum sensing and increased competence-induced killing or fratricide. These findings reveal a novel, previously unsuspected mechanism by which certain PPS-specific antibodies exert a direct effect on pneumococcal biology that has broad implications for bacterial clearance, genetic exchange, and antibody immunity to pneumococcus.

scholarly journals Development and Validation of 13-plex Luminex-Based Assay for Measuring Human Serum Antibodies to Streptococcus pneumoniae Capsular Polysaccharides

mSphere ◽  
2018 ◽  
Vol 3 (4) ◽  
pp. e00128-18 ◽  
Author(s):  
Danka Pavliakova ◽  
Peter C. Giardina ◽  
Soraya Moghazeh ◽  
Shite Sebastian ◽  
Maya Koster ◽  
...  
Keyword(s):  
Human Serum ◽  
Lower Limit ◽  
Immunosorbent Assay ◽  
Enzyme Linked Immunosorbent Assay ◽  
Igg Antibodies ◽  
Igg Antibody ◽  
Correlate Of Protection ◽  
Immune Correlate ◽  
Relative Standard ◽  
Limit Of Quantitation

ABSTRACT A Luminex-based direct immunoassay (dLIA) platform has been developed to replace the standardized pneumococcal enzyme-linked immunosorbent assay platform. The multiplex dLIA simultaneously measures the concentration of serum immunoglobulin G (IgG) antibodies specific for pneumococcal capsular polysaccharide (PnPS) serotypes 1, 3, 4, 5, 6A, 6B, 7F, 9V, 14, 18C, 19A, 19F, and 23F. The assay uses poly-l-lysine (PLL)-conjugated PnPS, chemically coupled to spectrally distinct Luminex microspheres. Assay validation experiments were performed using residual human serum samples obtained from 13-valent pneumococcal conjugate vaccine (13vPnC) clinical studies. Assay results are expressed as IgG antibody concentrations in micrograms per milliliter using the international reference serum, 007sp. The lower limit of quantitation (LLOQ) for all serotypes covered in the 13-plex dLIA fell within the range of 0.002 to 0.038 µg/ml serum IgG. The difference between the lower limit and upper limit of the assay range was >500-fold for all serotypes, and assay variability was <20% relative standard deviation (RSD) for all serotypes. IgG antibody measurements were shown to be serotype-specific (some cross-reactivity was observed only between the structurally related serotypes 6A and 6B as well as 19A and 19F), and no interference was observed between the serotypes when the assay was performed in the 13-plex format compared to the singleplex assays. The 13-plex dLIA platform developed by Pfizer Inc. generates up to 143 test results in a single 96-well plate and is a suitable replacement of the enzyme-linked immunosorbent assay (ELISA) platform for evaluating vaccine clinical trials. IMPORTANCE The pneumococcal enzyme-linked immunosorbent assay (ELISA) measures IgG antibodies in human serum, and it is an important assay that supports licensure of pneumococcal vaccines. The immune correlate of protection, 0.35 µg/ml of IgG antibodies, was determined by the ELISA method. Pfizer has developed a new Luminex-based assay platform to replace the ELISA. These papers describe the important work of (i) validating the Luminex-based assay and (ii) bridging the immune correlate of protection (0.35 µg/ml IgG) to equivalent values reported by the Luminex platform.

Enzyme-linked immunosorbent assay for cholesteryl ester transfer protein in human serum

1995 ◽  
Vol 240 (1) ◽  
pp. 1-9 ◽  
Author(s):  
Toshitaka Sato ◽  
Masayoshi Fukasawa ◽  
Makoto Kinoshita ◽  
Hiroyuki Arai ◽  
Takao Saeki ◽  
...  
Keyword(s):  
Cholesteryl Ester ◽  
Human Serum ◽  
Immunosorbent Assay ◽  
Cholesteryl Ester Transfer Protein ◽  
Enzyme Linked Immunosorbent Assay ◽  
Transfer Protein
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