Typing of HLA class II and class I antigens using PHA-activated, IL-2-propagated T lymphocytes.
We describe here a simple procedure, by which HLA class II antigens can be accurately and reliably identified in those patients where there is minimal or absent expression of HLA-DR,DQw antigens on B cells, or when the total number of leukocytes recovered from the patients do not permit reliable typing. Ficoll-Hypaque-separated peripheral blood mononuclear leukocytes, fresh or cryopreserved, were activated by PHA and then propagated in IL-2-containing medium until enough cells for typing were obtained (usually 7-14 days). At this stage, the cultured cells were shown to be primarily T cells (greater than 90% CD3+). Since the activated T cells propagate in the presence of IL-2, even a small number (10(4] of fresh or cryopreserved patients' cells suffice for this protocol. To date we have been able to successfully HLA-DR,DQw type 34/34 bone marrow transplantation candidates and 12/12 long-term dialysis patients, who were untypable using fresh cells. HLA-DR,DQw antigens on activated T cells from normal individuals were identical to those found on their uncultured B cells. In addition, class I antigens that were undetectable on the uncultured cells of one patient could be identified on activated T cells. The HLA antigens identified on the patients' activated T cells were confirmed by phenotypic analysis of cells from family members.
Leshem
B
B
Lautenberg Center for General and Tumor Immunology, Hebrew University, Jerusalem, Israel.
Cohen
I
I
Sherman
L
L
Brautbar
C
C
Kedar
E
E
D001402:B-Lymphocytes / Q000032:analysis; D002469:Cell Separation; D005260:Female; D006680:HLA Antigens / Q000032:analysis; D006681:HLA-D Antigens / Q000032:analysis; D006650:Histocompatibility Testing / Q000379:methods*; D006801:Humans; D007376:Interleukin-2 / Q000494:pharmacology*; D008213:Lymphocyte Activation; D008297:Male; D010641:Phenotype; D010835:Phytohemagglutinins; D013601:T-Lymphocytes / Q000032:analysis* / Q000276:immunology
D006680:HLA Antigens; D006681:HLA-D Antigens; D007376:Interleukin-2; D010835:Phytohemagglutinins