Expression cloning of human B cell immunoglobulins

Area of Science:

• Immunology
• Oncology
• Molecular Biology

Background:

• Most lymphomas arise from B cells, with antigen selection implicated in their development.
• The antibody specificities of lymphoma cells and their role in lymphomagenesis are poorly understood.

Purpose of the Study:

• To develop and describe a strategy for characterizing antibody reactivity in human B cells.
• To enable unbiased, single-cell level analysis of the human antibody repertoire.

Main Methods:

• Flow cytometric isolation of single human B cells.
• RT-PCR amplification of immunoglobulin heavy (Igh), kappa (Igκ), and lambda (Igλ) chain genes.
• Cloning into Ig gene expression vectors for in vitro production of recombinant monoclonal antibodies.

Main Results:

• A detailed protocol for generating recombinant monoclonal antibodies from single primary human B cells.
• The method allows for unbiased characterization of antibody repertoire on a single-cell level.

Conclusions:

• This strategy provides a means to investigate antibody reactivity in human lymphoma B cells.
• It can yield insights into the clonal evolution of B cell lymphomas through single-cell Ig gene sequencing.