Antigen-specific B cell analyses

The characterization of antigen-specific B cell responses and formation of antibody secreting cells (plasmablasts, plasma cells) is important when treatment aims to induce antigen-specific antibodies (e.g. vaccination studies). Alternatively, treatment may intent to prevent antibody formation (prevention of alloimmunsiation, graft vs host-disease, inhibitory antibody formation against biologicals) or to achieve reduction of specific antibodies in cases where antibodies contribute to pathology (e.g. autoimmune diseases). Sanquin offers several assays to analyze the characteristics of the antigen-specific B cell compartment and unravel the specific antibody repertoire in longitudinal analyses before and during treatment.

Antigen-specific B cell responses can either be analyzed by: 

  • Assessment of antigen-specific and Ig subclass-specific B cell frequencies by stimulating PBMCs in limiting dilution and analysing of the antibody titer and subclass specificity in ELISA (Pinna D et al. Eur J Immunol 2009; 39(5):1260-70).
 
  • Assessment of antigen-specific B cell frequencies by ELIspot (currently in development)
 
  • In vitro antigen-specific B cell proliferation upon CD4 T cell dependent or CD4 T cell independent B cell stimulation using a proliferation dye. This assay can be combined with Ig subclass specific antibodies to assess occurrence of division-linked isotype switching.
 
  • Isolation of antigen-specific B cells using fluorescent labelled antigen and FACS sort, followed by limiting dilution assays and ELISAs for antigen-specific antibodies. Specific VH/VL genes of identified antigen-specific B cells can be amplified and antigen-specific recombinant antibodies can be expressed in either HEK293 or CHO cells
 
  • Specific antibodies can be analyzed for titer, subclass specificity, affinity/avidity and specificity of target binding (see also under serology and immunogenicity analysis).

Assessment of specific CD4 populations that correlate to efficacy of induction of humoral immunity can be performed via extended ex vivo immunophenotyping of the CD4 T cell compartment before and during treatment (Cellular immunophenotyping)

Also see:

References

van Schouwenburg PA, Kruithof S, Votsmeier C, van Schie K, Hart MH, de Jong RN, van Buren EE, van Ham M, Aarden L, Wolbink G, Wouters D, Rispens T. Functional analysis of the anti-adalimumab response using patient-derived monoclonal antibodies. J Biol Chem 2014; 289(50):34482-8.

van Schouwenburg PA, van de Stadt LA, de Jong RN, van Buren EE, Kruithof S, de Groot E, Hart M, van Ham SM, Rispens T, Aarden L, Wolbink GJ, Wouters D. Adalimumab elicits a restricted anti-idiotypic antibody response in autoimmune patients resulting in functional neutralisation. Ann Rheum Dis 2013; 72(1):104-9.

de Wit J, Jorritsma T, Makuch M, Remmerswaal EB, Klaasse Bos H, Souwer Y, Neefjes J, ten Berge IJ, van Ham SM. Human B cells promote T-cell plasticity to optimize antibody response by inducing coexpression of T(H)1/T(FH) signatures. J Allergy Clin Immunol 2015; 135(4):1053-60.

Lighaam LC, Vermeulen E, Bleker Td, Meijlink KJ, Aalberse RC, Barnes E, Culver EL, van Ham SM, Rispens T. Phenotypic differences between IgG4+ and IgG1+ B cells point to distinct regulation of the IgG4 response. J Allergy Clin Immunol 2014; 133(1):267-70.

Last edited on: 4 November 2015