A primary study on establishing T-cell clones in vitro from bone marrow of a severe aplastic anemia patient / 中国免疫学杂志

ABSTRACT

Objective:To isolate and establish long term T cell clones derived from bone marrow(BM) of a patient with severe aplastic anemia(sAA).Methods:CD34 + cells and CD3 +T cells were enriched from BM aspirates,respectively by positive and negative immunomagnetic sorting protocol.BM of exicised rib of a non hematological patient was used as control. Irradiated CD34 + cells together with T cells were subject to one way autologous mixed lymphocyte culture(AMLC).Viable T cells surviving from AMLC were limit diluted and single cell was plated into microtitier wells along with feeder cells.Then all wells were screened for the growing cell wells and they were transferred into new wells to be expanded and maintained.Each clones obtained were primarily assessed CD4/CD8 phenotype and their homogeneity by immunocytochemical analysis.Results:The purity of sorted CD34 + and T cells was more than 90%. On AMLC,a total of 4?10 3 cells were viable with sAA,but 65 cells viable with control.Upon single cell seeding it 11 growing cell wells were found for sAA,and no one growing cells for control.The actual plating rate(2.1%) was within Poisson theoretic value range (