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Development and validation of a multiplex immunoassay for the simultaneous determination of serum antibodies to Bordetella pertussis, diphtheria and tetanus.
J Immunol Methods. 2008 Jun 01; 335(1-2):79-89.JI

Abstract

To increase testing of vaccine induced humoral immunity in immune surveillance studies and vaccine trials, a rapid and simple microsphere-based multiplex assay (pentaplex) was developed for the quantitation of IgG serum antibodies directed against the Bordetella pertussis antigens: Pertussis Toxin (Ptx), Filamentous hemagglutinin (FHA), Pertactin (Prn) and to Diphtheria toxin and Tetanus toxin. All individual antigens were covalently linked to carboxylated microspheres. The method was validated with different serum panels (n=60-78 samples). With the Multiplex Immunoassay (MIA) no evidence for bead interference between monoplex and pentaplex was found. The specificity of the method was shown by a heterologous inhibition of <16% and homologous inhibition of >92%. The pentaplex MIA appeared sensitive with lower limits of quantitation (LLOQ) well below those for ELISA (enzyme-linked immuno-sorbant assay). Assay reproducibility was high with intra-assay variability less than 10% and inter-assay variability below 14%. The reproducibility of the bead conjugation was good and beads could be stored up to at least 6 months without quality reduction. Importantly, the correlation of the pentaplex MIA with the individual ELISAs was excellent, R>0.98 for the Pertussis antigens and R=0.95 for Diphtheria and R=0.98 for Tetanus. Serum IgG antibodies to B. pertussis, Diphtheria and Tetanus can be measured easily, specific and reproducible using the pentaplex MIA. The pentaplex MIA shares features of the ELISA with the additional advantages of high sample throughput and small sample volumes and antigen required.

Authors+Show Affiliations

Laboratory for Infectious Diseases and Perinatal Screening, National Institute of Public Health and the Environment, Bilthoven, The Netherlands. pieter.van.gageldonk@rivm.nlNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Comparative Study
Journal Article
Validation Study

Language

eng

PubMed ID

18407287

Citation

van Gageldonk, Pieter G M., et al. "Development and Validation of a Multiplex Immunoassay for the Simultaneous Determination of Serum Antibodies to Bordetella Pertussis, Diphtheria and Tetanus." Journal of Immunological Methods, vol. 335, no. 1-2, 2008, pp. 79-89.
van Gageldonk PG, van Schaijk FG, van der Klis FR, et al. Development and validation of a multiplex immunoassay for the simultaneous determination of serum antibodies to Bordetella pertussis, diphtheria and tetanus. J Immunol Methods. 2008;335(1-2):79-89.
van Gageldonk, P. G., van Schaijk, F. G., van der Klis, F. R., & Berbers, G. A. (2008). Development and validation of a multiplex immunoassay for the simultaneous determination of serum antibodies to Bordetella pertussis, diphtheria and tetanus. Journal of Immunological Methods, 335(1-2), 79-89. https://doi.org/10.1016/j.jim.2008.02.018
van Gageldonk PG, et al. Development and Validation of a Multiplex Immunoassay for the Simultaneous Determination of Serum Antibodies to Bordetella Pertussis, Diphtheria and Tetanus. J Immunol Methods. 2008 Jun 1;335(1-2):79-89. PubMed PMID: 18407287.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Development and validation of a multiplex immunoassay for the simultaneous determination of serum antibodies to Bordetella pertussis, diphtheria and tetanus. AU - van Gageldonk,Pieter G M, AU - van Schaijk,Frank G, AU - van der Klis,Fiona R, AU - Berbers,Guy A M, Y1 - 2008/03/26/ PY - 2007/12/14/received PY - 2008/02/27/revised PY - 2008/02/28/accepted PY - 2008/4/15/pubmed PY - 2008/7/10/medline PY - 2008/4/15/entrez SP - 79 EP - 89 JF - Journal of immunological methods JO - J Immunol Methods VL - 335 IS - 1-2 N2 - To increase testing of vaccine induced humoral immunity in immune surveillance studies and vaccine trials, a rapid and simple microsphere-based multiplex assay (pentaplex) was developed for the quantitation of IgG serum antibodies directed against the Bordetella pertussis antigens: Pertussis Toxin (Ptx), Filamentous hemagglutinin (FHA), Pertactin (Prn) and to Diphtheria toxin and Tetanus toxin. All individual antigens were covalently linked to carboxylated microspheres. The method was validated with different serum panels (n=60-78 samples). With the Multiplex Immunoassay (MIA) no evidence for bead interference between monoplex and pentaplex was found. The specificity of the method was shown by a heterologous inhibition of <16% and homologous inhibition of >92%. The pentaplex MIA appeared sensitive with lower limits of quantitation (LLOQ) well below those for ELISA (enzyme-linked immuno-sorbant assay). Assay reproducibility was high with intra-assay variability less than 10% and inter-assay variability below 14%. The reproducibility of the bead conjugation was good and beads could be stored up to at least 6 months without quality reduction. Importantly, the correlation of the pentaplex MIA with the individual ELISAs was excellent, R>0.98 for the Pertussis antigens and R=0.95 for Diphtheria and R=0.98 for Tetanus. Serum IgG antibodies to B. pertussis, Diphtheria and Tetanus can be measured easily, specific and reproducible using the pentaplex MIA. The pentaplex MIA shares features of the ELISA with the additional advantages of high sample throughput and small sample volumes and antigen required. SN - 0022-1759 UR - https://www.unboundmedicine.com/medline/citation/18407287/Development_and_validation_of_a_multiplex_immunoassay_for_the_simultaneous_determination_of_serum_antibodies_to_Bordetella_pertussis_diphtheria_and_tetanus_ DB - PRIME DP - Unbound Medicine ER -