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Monitoring antigen-specific memory B cells and the antibodies they encode is important for understanding the specificity, breadth and duration of immune response to an infection or vaccination. The antibodies isolated could further help design vaccine antigens for raising relevant protective immune responses. However, developing assays to measure and isolate antigen-specific memory B cells is technically challenging due to the low frequencies of these cells that exist in the circulating blood. Here, we describe a flow cytometry method to identify and isolate Dengue envelope-specific memory B cells using a labelled Dengue envelope protein. We enumerated dengue-envelope specific memory B cells from a cohort of dengue seropositive donors using this direct flow cytometry assay. A more established and conventional assay, the cultured B ELISPOT, was used as a benchmark comparator. Furthermore, we were able to confirm the single-sorted memory B-cell specificity by culturing B cells and differentiating them into plasma cells using cell lines expressing CD40L. The culture supernatants were assayed for antigen binding and the ability of the antibodies to neutralize the cognate dengue virus. Moreover, we successfully isolated the heavy and light Ig sequences and expressed them as full-length recombinant antibodies to reproduce the activity seen in culture supernatants. Mapping of these antibodies revealed a novel epitope for dengue 2 virus serotype. In conclusion, we established a reproducible methodology to enumerate antigen-specific memory B cells and assay their encoded antibodies for functional characterization.
This article was published in the following journal.
Rapid diagnosis tests (RDTs) are easy to carry out, provide fast results, and could potentially guide medical treatment decisions. We investigated the performance of a commercially available RDT, whic...
Dengue is a globally important disease caused by four serotypes of dengue virus. Dengue vaccine development has been hampered by antigenic cross-reactivity among serotypes, which potentially causes an...
The Indian population is facing highest dengue burden worldwide supporting an urgent need for vaccines. For vaccine introduction, evaluation and interpretation it is important to gain a critical under...
Vaccination for dengue with the live attenuated tetravalent CYD-TDV vaccine (Dengvaxia®) is only recommended in individuals who have had prior dengue virus (DENV) infection. Rapid diagnostic tests (R...
Broadly neutralizing antibodies (bNAbs) are rarely elicited by current human immunodeficiency virus type 1 (HIV-1) vaccine designs, but the presence of bNAbs in naturally infected individuals may be a...
The purpose of this study is to assess the neutralizing antibody response against each dengue serotype post-vaccination.
The purpose of this study is to describe the neutralizing antibody response against each dengue serotype at 1 month post second dose of TDV or placebo in dengue-naive adolescent participan...
This is a follow up study using three different formulations for each serotype with the aim of testing immunogenicity and reactogenicity Primary Objective: Immunogenicity: To evalu...
In natural HIV disease, a small fraction (1-2%) of infected individuals develops exceptionally high titres of HIV-1 neutralizing serum activity. Antibodies isolated from these individuals ...
The aim of the study is to assess and describe the booster effect of a CYD dengue vaccine dose administered 4 to 5 years after the completion of a 3-dose vaccination schedule. Primary Obj...
An acute infectious, eruptive, febrile disease caused by four antigenically related but distinct serotypes of the DENGUE VIRUS. It is transmitted by the bite of infected Aedes mosquitoes, especially A. aegypti. Classical dengue (dengue fever) is self-limiting and characterized by fever, myalgia, headache, and rash. DENGUE HEMORRHAGIC FEVER is a more virulent form of dengue virus infection and a separate clinical entity. (From Dorland, 28th ed)
Antibodies that reduce or abolish some biological activity of a soluble antigen or infectious agent, usually a virus.
Development of neutralizing antibodies in individuals who have been exposed to the human immunodeficiency virus (HIV/HTLV-III/LAV).
A distinct and virulent form of DENGUE characterized by thrombocytopenia and hemoconcentration (grades I and II) and distinguished by a positive tourniquet test. When accompanied by circulatory failure and shock (grades III and IV), it is called dengue shock syndrome. (From Dorland, 28th ed)
A virulent form of dengue characterized by THROMBOCYTOPENIA and an increase in vascular permeability (grades I and II) and distinguished by a positive pain test (e.g., TOURNIQUET PAIN TEST). When accompanied by SHOCK (grades III and IV), it is called dengue shock syndrome.
Tropical Medicine is the study of diseases more commonly found in tropical regions than elsewhere. Examples of these diseases are malaria, yellow fever, Chagas disease, Dengue, Helminths, African trypanosomiasis, Leishmaniasis, Leprosy, Lymphatic filaria...
An antibody is a protein produced by the body's immune system when it detects harmful substances, called antigens. Examples of antigens include microorganisms (such as bacteria, fungi, parasites, and viruses) and chemicals. Antibodies may be produc...