Improved biochemical assay for terminal deoxynucleotidyl transferase in human blood cells: Results in 89 adult patients with lymphoid leukemias and malignant lymphomas in leukemic phase

Ion filtration chromatography of crude cell homogenates on DEAE-Sephadex allows determination of terminal deoxynucleotidyl transferase (TdT) activity in 10⁶ blast cells from acute lymphoblastic leukemia (ALL) and in 2 × 10⁷ normal bone marrow (BM) cells. Ninety-seven determinations of TdT in 40 patients with ALL during various stages of their disease revealed high levels of activity in BM and peripheral blood samples from all patients studied at diagnosis and in relapse. In $\text{10}\text{34}$ BM samples from patients with ALL in remission, levels of TdT activity were found to be significantly elevated as compared to normal controls. The remaining cases exhibited TdT activities within the normal range. In 20 patients with non-Hodgkin's lymphomas of null and T cell type in leukemic phase, TdT activities were within the same range as observed in active ALL. Eighteen of these had a histological diagnosis of diffuse poorly differentiated lymphoma of lymphoblastic type, one of giant follicular lymphoma and one of diffuse histiocytic lymphoma. In two patients with unclassifiable lymphoproliferative diseases of T cell type, no TdT activity was found, possibly indicating a disease of mature T cells. All 27 patients with lymphoid neoplasias of B cell type were found to exhibit no TdT activity in involved tissues. Determination of TdT activity appears to be a sensitive assay for detection of subclinical bone marrow involvement in TdT positive lymphoproliferative diseases. The clinical and theoretical significance of these observations is discussed.     

Preparation and characterization of monoclonal antibodies recognizing two distinct differentiation antigens (Pro-Im1, Pro-Im2) on early hematopoietic cells

A series of monoclonal antibodies recognizing myeloid differentiation antigens were prepared by immunizing Balb/c mice with HL-60 cells. Hybrids secreting antibodies reactive with HL-60 cells but unreactive with peripheral blood mononuclear cells were isolated and further cloned. One clone was found to produce an IgG2a antibody recognizing an 85,000-dalton molecular weight surface glycoprotein, and a second clone was found to produce an IgM antibody recognizing a heat-stable determinant present on a glycolipid. We have termed these antigens Pro- Im1 and Pro-Im2, respectively (Pro for using HL-60 promyelocytes as an immunogen and Im for the presence of these antigens on immature cells). alpha Pro-Im1 and alpha Pro-Im2 were used to investigate the surface expression and tissue distribution of these two antigens. Pro-Im1 and Pro-Im2 were found to be brightly expressed on a fraction of fetal liver hematopoietic and bone marrow cells. Both antibodies mediated complement-dependent inhibition of CFU-GM, BFU-E, and CFU-GEMM formation assayed by soft agar colony and burst formation, indicating the expression of these antigens by early hematopoietic precursor cells. This was further confirmed by the induction of HL-60 cells by TPA to differentiate into more mature monocytes and macrophages, accompanied by the loss of both antigens. Pro-Im1 and Pro-Im2 were absent from peripheral blood monocytes, erythrocytes, and platelets, but Pro-Im2 was expressed on granulocytes. Both antigens were absent from thymocytes and peripheral T cells. Cytofluorographic analysis suggested their absence from peripheral blood B cells but that both were expressed on a minority of tissue B cells. Analysis of 150 cases of various myeloid and lymphoid malignancies demonstrated Pro-Im1 and Pro-Im2 expression on myeloblasts and promyelocytes from some acute myelogenous leukemias as well as some B cell malignancies, suggesting that these antigens are shared by early hematopoietic cells and a subset of B cells.