Development and Validation of 13-plex Luminex-Based Assay for Measuring Human Serum Antibodies to <italic toggle="yes">Streptococcus pneumoniae</italic> Capsular Polysaccharides

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 capsul...

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Autores principales:	Danka Pavliakova, Peter C. Giardina, Soraya Moghazeh, Shite Sebastian, Maya Koster, Viliam Pavliak, Andrew McKeen, Roger French, Kathrin U. Jansen, Michael Pride
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2018
Materias:	Luminex development multiplex pneumococcal validation Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/dbf3807ed7cc4c2999a7c6d1253b02f6
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spelling	oai:doaj.org-article:dbf3807ed7cc4c2999a7c6d1253b02f62021-11-15T15:25:50ZDevelopment and Validation of 13-plex Luminex-Based Assay for Measuring Human Serum Antibodies to <italic toggle="yes">Streptococcus pneumoniae</italic> Capsular Polysaccharides10.1128/mSphere.00128-182379-5042https://doaj.org/article/dbf3807ed7cc4c2999a7c6d1253b02f62018-08-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mSphere.00128-18https://doaj.org/toc/2379-5042ABSTRACT 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.Danka PavliakovaPeter C. GiardinaSoraya MoghazehShite SebastianMaya KosterViliam PavliakAndrew McKeenRoger FrenchKathrin U. JansenMichael PrideAmerican Society for MicrobiologyarticleLuminexdevelopmentmultiplexpneumococcalvalidationMicrobiologyQR1-502ENmSphere, Vol 3, Iss 4 (2018)
institution	DOAJ
collection	DOAJ
language	EN
topic	Luminex development multiplex pneumococcal validation Microbiology QR1-502
spellingShingle	Luminex development multiplex pneumococcal validation Microbiology QR1-502 Danka Pavliakova Peter C. Giardina Soraya Moghazeh Shite Sebastian Maya Koster Viliam Pavliak Andrew McKeen Roger French Kathrin U. Jansen Michael Pride Development and Validation of 13-plex Luminex-Based Assay for Measuring Human Serum Antibodies to <italic toggle="yes">Streptococcus pneumoniae</italic> Capsular Polysaccharides
description	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.
format	article
author	Danka Pavliakova Peter C. Giardina Soraya Moghazeh Shite Sebastian Maya Koster Viliam Pavliak Andrew McKeen Roger French Kathrin U. Jansen Michael Pride
author_facet	Danka Pavliakova Peter C. Giardina Soraya Moghazeh Shite Sebastian Maya Koster Viliam Pavliak Andrew McKeen Roger French Kathrin U. Jansen Michael Pride
author_sort	Danka Pavliakova
title	Development and Validation of 13-plex Luminex-Based Assay for Measuring Human Serum Antibodies to <italic toggle="yes">Streptococcus pneumoniae</italic> Capsular Polysaccharides
title_short	Development and Validation of 13-plex Luminex-Based Assay for Measuring Human Serum Antibodies to <italic toggle="yes">Streptococcus pneumoniae</italic> Capsular Polysaccharides
title_full	Development and Validation of 13-plex Luminex-Based Assay for Measuring Human Serum Antibodies to <italic toggle="yes">Streptococcus pneumoniae</italic> Capsular Polysaccharides
title_fullStr	Development and Validation of 13-plex Luminex-Based Assay for Measuring Human Serum Antibodies to <italic toggle="yes">Streptococcus pneumoniae</italic> Capsular Polysaccharides
title_full_unstemmed	Development and Validation of 13-plex Luminex-Based Assay for Measuring Human Serum Antibodies to <italic toggle="yes">Streptococcus pneumoniae</italic> Capsular Polysaccharides
title_sort	development and validation of 13-plex luminex-based assay for measuring human serum antibodies to <italic toggle="yes">streptococcus pneumoniae</italic> capsular polysaccharides
publisher	American Society for Microbiology
publishDate	2018
url	https://doaj.org/article/dbf3807ed7cc4c2999a7c6d1253b02f6
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Development and Validation of 13-plex Luminex-Based Assay for Measuring Human Serum Antibodies to <italic toggle="yes">Streptococcus pneumoniae</italic> Capsular Polysaccharides

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