Comparison of non-magnetic and magnetic beads in bead-based assays.
Multiplex bead-based flow cytometry is an attractive way for simultaneous, rapid and cost-effective analysis of multiple analytes in a single sample. Previously, we developed various bead-based assays using non-magnetic beads coated with Staphylococcus aureus and Streptococcus pneumoniae antigens for the detection of antibodies. Here, we compared the performance of the assay using non-magnetic beads with one based on the newly developed magnetic beads. We optimized the magnetic beads' coupling procedure and antibody detection assays for S. aureus and S. pneumoniae antigens and we measured IgG in human pooled serum against a series of S. aureus and S. pneumoniae-derived antigens in a singleplex and in a multiplex assay, respectively. For the multiplex assay, the comparison between magnetic and non-magnetic beads showed: i) in the majority of the cases (13 of the 17 tested S. pneumoniae antigens) significantly higher Median Fluorescence Intensity (MFI) values, ii) lower detection limits, iii) lower coefficient of variation (CV: 12% vs. 7% for non-magnetic vs. magnetic beads), so lower inter-assay variation and hence higher reproducibility. Magnetic bead coupling is cost effective, as we used 25% of the normal amount of antigen and only 50% of the beads in comparison to the non-magnetic beads. This optimized magnetic-based assay, which combines ease of use with an improved assay performance, allows detection of antibodies with a low titer that are potentially missed with the non-magnetic-based assay.
Department of Medical Microbiology and Infectious Diseases, Erasmus MC, Rotterdam, The Netherlands; Department of Oral Biochemistry, Academic Centre for Dentistry Amsterdam (ACTA), University of Amsterdam and VU University Amsterdam, Amsterdam, The Netherlands. Electronic address: s.hansenovamanaskova@erasmusmc.nl.
Silvie
S
Hansenová Maňásková
Chief Scientific Office, BioMérieux, La Balme les Grottes, France.
Alex
A
van Belkum
Department of Medical Microbiology and Infectious Diseases, Erasmus MC, Rotterdam, The Netherlands; International Centre for Diarrhoeal Disease Research Bangladesh, Dhaka, Bangladesh; Fondation Mérieux, Lyon, France.
Hubert P
HP
Endtz
Department of Oral Biochemistry, Academic Centre for Dentistry Amsterdam (ACTA), University of Amsterdam and VU University Amsterdam, Amsterdam, The Netherlands.
Floris J
FJ
Bikker
Department of Oral Biochemistry, Academic Centre for Dentistry Amsterdam (ACTA), University of Amsterdam and VU University Amsterdam, Amsterdam, The Netherlands.
Enno C I
EC
Veerman
Department of Medical Microbiology and Infectious Diseases, Erasmus MC, Rotterdam, The Netherlands.
Willem J B
WJ
van Wamel
D000942:Antigens, Bacterial / Q000276:immunology*; D005434:Flow Cytometry / Q000379:methods*; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood*; D018189:Immunomagnetic Separation / Q000191:economics / Q000379:methods*; D008863:Microspheres; D015203:Reproducibility of Results; D013211:Staphylococcus aureus; D013296:Streptococcus pneumoniae
D000942:Antigens, Bacterial; D007074:Immunoglobulin G
Antigen coupling to magnetic beads; Bead-based assay; Microspheres; Staphylococcus aureus; Streptococcus pneumoniae