Applications for B cell ELISpot
B cell ELISpot is an assay relevant wherever it is essential to characterize an antibody response. Antibody (humoral) responses are generated by B cells (part of the adaptive immune system) against foreign agents, and B cell ELISpots are a way of specifically measuring these responses.
For the most part antibody responses are useful, and lead to the neutralization of pathogens such as influenza virus. Vaccine developers depend on being able to elicit an antibody response using a carefully designed vaccine; this response will then be protective against future encounters with a particular pathogen. B cell ELISpot is thus useful at two points in vaccine development – firstly in characterizing the normal antibody response to a pathogen, and then later in measuring the protective antibody response that a vaccine can elicit in patients.
Examples of unwanted antibody responses include responses directed against novel protein therapeutics, which can render the therapy ineffective or even cause harm to a patient, and the autoantibodies produced in autoimmune disease. B cell ELISpot presents an excellent way to investigate both of these. In addition, the effects of immune modulation - by drugs, or as a consequence of immune deficiency - can also be monitored precisely using B cell ELISpot.
Influenza presents a health risk to vulnerable individuals in the guise of ‘seasonal flu’, and to the general population as strains of highly contagious flu continue to evolve 1. It is important to understand the best vaccination strategy for eliciting a protective response – although vaccines may be tweaked in response to each strain of flu, a reliable method of eliciting a response from a vaccine is necessary. Many research teams have relied on B cell ELISpot to inform them of when their vaccines are working. Examples of these experiments include the work of Joo et al 2, who used B cell ELISpot to compare the antibody-secreting cells produced in response to different vaccination sites, Halliley et al 3 , who investigated when the post-vaccine peak of antibody-producing cells occurred, Baer et al 4, who analyzed responses to different ‘flu strains to quantify antibody cross-reactivity, and by researchers investigating the existing polyclonal anti-influenza response in humans seeking to generate anti-flu vaccines with broad specificity 5. Malaria is another example of a highly infectious disease and a global health problem, and the antibody responses made against Plasmodium parasites (the causative agent for malaria) can be efficiently measured using B cell ELISpot 6.
Vaccination against childhood diseases has been established as a reliable way to decrease their incidence. Mumps, Whooping cough and Meningitis researchers have all made use of the power of B cell ELISpot to monitor patient responses to disease and to novel vaccines 7-10.
In HIV research, too, B cell ELISpot has been used to great effect in characterizing the humoral response to virus. The B cell response to HIV-1 envelope glycoprotein (Env) has been described in detail using this technique 11, as have the pool of memory B cells elicited by a prime/boost vaccination strategy 12, and the loss of humoral responses to common childhood vaccination antigens in HIV patients presenting for anti-retroviral treatment 13.
Vaccination strategies to combat cancer have been heavily investigated over the last decade, and can be classed as anti-tumor or preventative (where a tumor has a known viral cause) vaccines. For example, vaccination against human papillomavirus (HPV) is being used in an attempt to reduce the incidence of cervical cancer, and ongoing work will be necessary to quantify the duration of a protective antibody response against HPV elicited by Gardasil® (Merck) and Cervarix® (GSK) 14. B cell ELISpot would be ideal for this work. Recent work with a therapy designed to target tumor vascularization, and so starve tumors and prevent them from growing, used B cell ELISpot to correlate the antibody response induced by treatment with the effects on tumor vasculature 15.
Hemorrhagic fevers caused by the Ebola virus family (filoviruses) are little-studied because of their lethal nature, but represent a possible bioweapon. As such, labs at the United States Army Medical Research Institute of Infectious Diseases are investigating them to see if there might be a possible treatment. As part of their work they have use B cell ELISpot to characterize responses to filovirus 16.
Autoimmunity and Allergy
IgE antibodies are a central part of the development of allergic conditions, and assessment of the ability of individuals to mount an IgE response in different conditions has been carried out using B cell ELISpot 17. It is critical to understand the effects of different settings on a humoral response (for example the presence of immunomodulatory agents, immunosuppression caused by infection or chemotherapy) and B cell ELISpot is an excellent assay to apply to these questions.
As mentioned above, unwanted antibody responses are at the core of autoimmune diseases. B cell ELISpot has been used extensively in autoimmunity research, both in basic research into the development of autoimmune responses 18, 19, and to explore responses in patients undergoing treatment 20, 21.
Shigella infection can lead to dysentery, so vaccination with a live attenuated version has been investigated as a preventative measure. B cell ELISpot was used as a tool to monitor Shigella antigen-specific antibody responses in infection 22, and also to verify the immunogenicity generated by an attenuated version 23.
(2) Joo et al Quantitative analysis of influenza virus-specific B cell memory generated by different routes of inactivated virus vaccination. Vaccine (2010) 28:2186 [PubMedID:20056191]
(3) Halliley et al Peak frequencies of circulating human influenza-specific antibody secreting cells correlate with serum antibody response after immunization. Vaccine (2010) 28:3582 [PubMedID:20298818]
(4) Baer et al B cell responses to H5 influenza HA in human subjects vaccinated with a drifted variant. Vaccine. (2010) 28:907 [PubMedID:19932673]
(5) Wrammert et al Rapid cloning of high-affinity human monoclonal antibodies against influenza virus. Nature (2008) 453:667 [PubMedID:18449194]
(6) Weiss et al High efficiency human memory B cell assay and its application to studying Plasmodium falciparum-specific memory B cells in natural infections. J Immunol Methods. (2012) 375:68. [PubMedID:21963949]
(7) Cruz et al Comparison of long-term humoral memory development after immunisation against Neisseria meningitidis B or diphtheria toxoid. Vaccine. (2010) 28:6841. [PubMedID:20719254]
(8) Cruz et al Generation of long-lived plasma cells to serogroup B Neisseria meningitidis after murine immunisation with an outer membrane protein vaccine. Vaccine. (2007) 25:5046 [PubMedID:17524531]
(9) Latner et al Enzyme-linked immunospot assay detection of mumps-specific antibody-secreting B cells as an alternative method of laboratory diagnosis. Clin Vaccine Immunol. (2011) 18:35 [PubMedID:21047998]
(10) George-Chandy et al Vaccination with Bordetella pertussis-pulsed autologous or heterologous dendritic cells induces a mucosal antibody response in vivo and protects against infection. Infect Immun. 69:4120 [PubMedID:11349085]
(11) Dosenovic et al Selective expansion of HIV-1 envelope glycoprotein-specific B cell subsets recognizing distinct structural elements following immunization. J Immunol. (2009) 183:3373 [PubMedID:19696434]
(12) Brocca-Cofano et al Vaccine-elicited SIV and HIV envelope-specific IgA and IgG memory B cells in rhesus macaque peripheral blood correlate with functional antibody responses and reduced viremia. Vaccine. (2011) 29:3310 21382487 [PubMedID:21382487]
(13) Pensieroso et al Timing of HAART defines the integrity of memory B cells and the longevity of humoral responses in HIV-1 vertically-infected children. Proc Natl Acad Sci U S A. (2009) 106:7939 [PubMedID:19416836]
(14) Romanowski B. Long term protection against cervical infection with the human papillomavirus: review of currently available vaccines. Hum Vaccin. (2011) 7:161 [PubMedID:21307652]
(15) Ren et al Inhibition of tumor angiogenesis in lung cancer by T4 phage surface displaying mVEGFR2 vaccine. Vaccine (2011) 29:5802. [PubMedID:21482223]
(16) Warfield K, Olinger G. Protective role of cytotoxic T lymphocytes in filovirus hemorrhagic fever. J Biomed Biotechnol. (2011) 2011:984241 [PubMedID:22253531]
(17) Esen et al Effect of IL-15 on IgG versus IgE antibody-secreting cells in vitro. J Immunol Methods. (2012) 375:7 [PubMedID:21945396]
(18) Ju et al Unexpected development of autoimmunity in BAFF-R-mutant MRL-lpr mice. Immunology. (2007) 120:281 [PubMedID:17073941]
(19) Enghard et al Subset size, activation threshold and distribution of autoreactive MZ and FO B cells do not differ in a sex-specific manner in the NZB/W F1 murine lupus model: an experimental mouse study. Lupus. (2011) 20:1240 [PubMedID:21844114]
(20) Jacobi et al Effect of long-term belimumab treatment on B cells in systemic lupus erythematosus: extension of a phase II, double-blind, placebo-controlled, dose-ranging study.Arthritis Rheum. (2010) 62:201 [PubMedID:20039404]
(21) Rehnberg et al Vaccination response to protein and carbohydrate antigens in patients with rheumatoid arthritis after rituximab treatment. Arthritis Res Ther. (2010) 12:R111. [PubMedID:20529331]
(22) Feller et al Comparative evaluation of the antibody in lymphocyte supernatant (ALS) and enzyme-linked immunospot (ELISPOT) assays for measuring mucosal immune responses to Shigella antigens. Vaccine. (2011) 29:8487 [PubMedID:21939714]
(23) McKenzie et al Safety and immunogenicity of WRSd1, a live attenuated Shigella dysenteriae type 1 vaccine candidate. Vaccine. (2008) 26:3291 [PubMedID:18468742]