Large-scale production of CD4+ T cells from HIV-1-infected donors after CD3/CD28 costimulation

J Hematother. 1998 Oct;7(5):437-48. doi: 10.1089/scd.1.1998.7.437.


We describe a procedure for large-scale enrichment, growth, and harvesting CD4+ T cells. This method may be effective for HIV-1 immunotherapy, as the mode of stimulation, with anti-CD3 plus anti-CD28 coated beads (CD3/CD28 beads) induces a potent antiviral effect. PBMC were obtained by density gradient centrifugation of an apheresis product. Monocytes/macrophages were removed by incubating PBMC with beads coated with IgG. The cells were then magnetically depleted of B cells and CD8+ cells with mouse anti-CD20 and anti-CD8 MAbs and sheep antimouse coated beads. The remaining cells were >80% CD4+ and were transferred to gas-permeable bags containing CD3/CD28 beads and cultured in a closed system. After 14 days, the cell number increased an average of 37-fold, and cells were nearly 100% CD4+. Viral load, assessed by DNA PCR for HIV-1 gag, decreased >10-fold during culture in the absence of antiretroviral agents. Removal of CD3/CD28 beads from the cell suspension was accomplished by passing cells plus beads (3-30 x 10(9) cells in 2-12 L) over a MaxSep magnetic separator using gravity-driven flow. The cells were then concentrated to 300 ml in an automated centrifuge. This process allows safe and efficient growth of large numbers of CD4+ T cells from HIV-1+ donors.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Adoptive Transfer / methods*
  • Animals
  • CD28 Antigens / immunology
  • CD3 Complex / immunology
  • CD4-Positive T-Lymphocytes / immunology
  • CD4-Positive T-Lymphocytes / pathology*
  • Cells, Cultured
  • Centrifugation, Density Gradient
  • HIV Infections / immunology
  • HIV Infections / therapy*
  • Humans
  • Immunosorbent Techniques
  • Leukapheresis / methods*
  • Mice


  • CD28 Antigens
  • CD3 Complex