3D Tissue Explant and Single-Cell Suspension Organoid Culture Systems for Ex Vivo Drug Testing on Human Tonsil-Derived T Follicular Helper Cells

Methods Mol Biol. 2022:2380:267-288. doi: 10.1007/978-1-0716-1736-6_22.


Research on the human immune system is often restricted to peripheral blood cells. However, these cells can be different from those found in secondary lymphoid organs. For instance, specialized T and B cells that are localized in germinal centers (GCs), which are complex anatomical structures being required for the generation of potent antibodies, are not found in peripheral blood. Most T helper cells located in GCs belong to the T follicular helper (Tfh) cell subset, which provides critical support to B cells. Bona fide human GC Tfh cells can be obtained from secondary lymphoid tissues such as tonsils, which are routinely removed by surgery. We here describe a method that is based on human lymphoid histoculture (HLH) and human lymphoid aggregate culture (HLAC) to culture human adenoid (pharyngeal tonsil) tissue ex vivo, followed by deep Tfh cell phenotyping by flow cytometry. This method allows studying Tfh cells in a versatile explant culture system that preserves many aspects of the original in vivo three-dimensional (3D) structure, in parallel to single-cell suspension organoid cultures in which the original tissue structure is disintegrated. We also describe how this versatile platform can be used for drug testing or manipulation of human Tfh cells in vitro for mechanistic studies.

Keywords: BCL6; CXCR5; Flow cytometry; Germinal center (GC); Human ex vivo lymphoid tissue culture; JAK inhibitor; Organoid; T follicular helper cells; Tfh; Tfh cell subsets; Tonsil.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Germinal Center
  • Humans
  • Organoids
  • Palatine Tonsil
  • Pharmaceutical Preparations
  • T Follicular Helper Cells*
  • T-Lymphocytes, Helper-Inducer
  • Tissue Culture Techniques


  • Pharmaceutical Preparations