Single Cell Analysis of Synovial Tissue B-Cells

  • Hye-Jung Kim
  • Claudia Berek
Part of the Methods in Molecular Medicine book series (MIMM, volume 136)

Abstract

Mononuclear cells often form highly organized lymphoid structures in the chronically inflamed synovial tissue of patients with rheumatoid arthritis (RA) within which B-cells are activated and may differentiate into effector plasma cells. The analysis of those activated B-cells and the determination of their specificity is of great importance for the understanding of the pathogenesis of RA. Here, we describe a technique that combines histological analysis of synovial tissue with a molecular analysis of the Vgene repertoire at the level of the single B-cell.

Immunohistochemical staining of tissue sections allows us to identify the activated B-cells. Those cells are then isolated using a micromanipulator and the rearranged immunoglobulin (Ig) genes amplified, cloned and sequenced. The combination of the V(D)J gene segments and the pattern of somatic mutations in the V-region genes, allows us to identify clonal relationships between the isolated B cells. Once Ig genes for a heavy and a light chain have been isolated from individual B-cells, they can be used to generate recombinant antibodies. These antibodies can be used to determine the antigens which support the activation of B-cells in the inflamed synovial tissue

Key Words

B-cells plasma cells immunoglobulin genes rheumatoid arthritis synovial tissue germinal center micromanipulation single cell PCR somatic hypermutation 

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Copyright information

© Humana Press Inc., Totowa, NJ 2007

Authors and Affiliations

  • Hye-Jung Kim
    • 1
  • Claudia Berek
    • 2
  1. 1.Memorial Sloan Kettering Cancer CenterDepartment of ImmunologyNew York
  2. 2.Deutsches RheumaForschungszentrumBerlinGermany

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