A quantitative flow cytometry assay for the preclinical testing and pharmacological monitoring of rabbit antilymphocyte globulins (rATG)

J Immunol Methods. 2000 Nov 1;245(1-2):45-54. doi: 10.1016/s0022-1759(00)00271-4.


Rabbit antilymphocyte or antithymocyte globulins (rATG) are currently used as immunosuppressive agents in clinical organ and bone marrow transplantation, but because of differences in antigen source and purification processes between manufacturers, determination of the dose to be administered remains empirical. Dosage may be adjusted to peripheral blood T cell counts or limited by side effects such as neutropenia or thrombocytopenia. We report here a measurement by indirect immunofluorescence of the amount of antibodies from rATG that bind to different blood cell types. A concentration-related immunofluorescence signal was obtained with reasonable replicability, allowing comparison of different rATGs by reference to the same rATG batch as internal standard. Ratios of antilymphocyte to undesirable antibodies (directed against antigens expressed on neutrophils, platelets or red cells) could be calculated for each rATG preparation. Finally, the method is applicable to the measurement of free antibodies in sera from patients treated with rATG and to the analysis of the effects of in vivo absorption of rATG during its administration. Marked differences in antibody content per mg of rIgG account for the differences in rATG dosage that were empirically established for clinical applications.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antilymphocyte Serum / analysis*
  • Antilymphocyte Serum / therapeutic use
  • B-Lymphocytes / immunology
  • Blood Platelets / immunology
  • CD4-Positive T-Lymphocytes / immunology
  • CD8-Positive T-Lymphocytes / immunology
  • Complement System Proteins / metabolism
  • Cytotoxicity Tests, Immunologic
  • Erythrocytes / immunology
  • Flow Cytometry / methods*
  • Fluorescent Antibody Technique, Indirect
  • Humans
  • Immunosuppressive Agents / analysis
  • Immunosuppressive Agents / therapeutic use
  • Killer Cells, Natural / immunology
  • Neutrophils / immunology
  • Rabbits
  • Transplantation Immunology


  • Antilymphocyte Serum
  • Immunosuppressive Agents
  • Complement System Proteins