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J Drach

Publications and source records attributed to J Drach.

At least 73 records · Page 4Linked to original sources
PubMed1994

Retinoic acid-induced expression of CD38 antigen in myeloid cells is mediated through retinoic acid receptor-alpha.

CD38 is a leukocyte differentiation antigen that has been thought to be a phenotypic marker of different subpopulations of T- and B-lymphocytes. In myeloid cells, CD38 is expressed during early stages of differentiation. Virtually no information is available on regulation and functions of CD38. Recently we reported that all-trans-retinoic acid (ATRA) is a potent and highly specific inducer of CD38 expression in human promyelocytic leukemia cells. Here we report that ATRA-induced expression of CD38 antigen in myeloid cells is mediated through retinoic acid-alpha receptor (RAR alpha). ATRA failed to induce CD38 expression in a mutant subclone of the HL-60 myeloid leukemia cell line (designated HL-60R) that is relatively resistant to ATRA-induced granulocytic differentiation. Retroviral vector-mediated transduction of RA receptor (RAR alpha) into this HL-60R subclone completely restored the sensitivity of these cells to ATRA in terms of their ability to express CD38. In contrast, CD38 expression was not inducible by ATRA in HL-60R cells, transfected with a functional RAR beta, RAR gamma, or RXR alpha receptor. Induction of CD38 in acute promyelocytic and acute myeloblastic leukemia cells was independent of ATRA-induced cytodifferentiation. Following culture with ATRA, increased CD38 protein levels were also observed in normal CD34+ bone marrow cells, but not on normal circulating granulocytes. From these results, we conclude that CD38 is ATRA inducible in myeloid leukemia cells and normal CD34+ bone marrow cells. This effect is independent of differentiation and is mediated by RAR alpha in HL-60 cells, suggesting a similar role for RAR alpha in CD38 expression in other hematopoietic cells.

ADP-ribosyl Cyclase↗
PubMed1994

Growth inhibition of follicular small-cleaved-cell lymphoma cells in short-term culture by interleukin-3.

BACKGROUND: The role of interleukin-3 (IL-3) in stimulating the growth of early myeloid progenitor cells is very well established. Therefore, IL-3 has been incorporated into many post-bone-marrow transplantation and intensive chemotherapy programs for the treatment of solid tumors and hematologic malignancies, including lymphomas. However, the effect of IL-3 on normal and malignant lymphocytes has not been well studied. PURPOSE: The purpose of this study was to evaluate the in vitro effect of IL-3 on the growth of follicular small-cleaved-cell lymphoma (FSCCL). MATERIALS AND METHODS: IL-3 receptor expression on the surface of CD19+ cells was determined by two-color flow cytometry measuring the receptor-binding of biotinylated IL-3 to CD19+ B-cells. Seven cases of FSCCL were compared to six normal controls. Cell proliferation was evaluated by [3H]thymidine incorporation into cells grown in suspension cultures. RESULTS: All seven cases of FSCCL expressed the IL-3 receptor on the surface of CD19+ cells, whereas all six cases of CD19+ cells isolated from the peripheral blood of normal donors did not express IL-3 receptors. IL-3 had antiproliferative activity against FSCCL as manifested by a decrease in [3H]thymidine incorporation and a decrease in the total number of cells after 72 hours of culture. CONCLUSION: IL-3 inhibits the growth of FSCCL cells in vitro. Clinical trials to evaluate the in vivo effect of IL-3 in patients with FSCCL are warranted.

Antigens, CD↗
PubMed1994

Granulocyte-colony stimulating factor, granulocyte-macrophage colony stimulating factor, PIXY-321, stem cell factor, interleukin-3, and interleukin-7: receptor binding and effects on clonogenic proliferation in acute lymphoblastic leukemia.

Cytokines are frequently used after chemotherapy of leukemias and solid tumors to augment recovery of normal hematopoiesis. While the regulation of normal and leukemic myelopoiesis is well investigated, little is known about effects of cytokines on growth and differentiation of lymphoblastic leukemia. In this study, we investigated the expression of receptors for G-CSF, GM-CSF, SCF, IL-3, and IL-7 on acute lymphoblastic leukemia (ALL) blasts and the effects of these growth factors (GF) on ALL blast colony formation. The binding of fluorescence-tagged cytokines to receptors on ALL blasts was studied by flow-cytometry in 27 cases of ALL (24 precursor B-ALL, 3 T-ALL). Receptor-binding for myeloid-associated GF was observed in the majority of precursor B-ALL (G-CSF = 100%, GM-CSF = 65%, IL-3 = 83%, SCF = 74%), but not in T-ALL. Binding of labelled IL-7 was detected in both precursor B- (92%) and T-ALL (100%). The presence of receptors for SCF in ALL was confirmed by polymerase chain reaction for c-kit mRNA in 19/21 cases tested. Expression of receptors for G-CSF, GM-CSF, IL-3, and SCF was not associated with expression of myeloid antigens, or with specific cytogenetic abnormalities. The effects of these GF on clonogenic cells were tested in the ALL blast colony assay and varied between samples, but all cytokines were able to increase clonogenic growth. The GM-CSF/IL-3 fusion molecule PIXY-321 was most effective in promoting colony growth. In some cases inhibition of colony formation was found. We conclude that ALL blast cells have receptors not only for IL-7, but also for G-CSF, GM-CSF, SCF, and IL-3. ALL precursors can respond to these GF with changes in their clonogenic growth indicating the presence of functional receptors. Results may have implications for therapeutic approaches combining cytokines and chemotherapy.

Adolescent↗
PubMed1994

Large-scale preparation of highly purified, frozen/thawed CD34+, HLA-DR- hematopoietic progenitor cells by sequential immunoadsorption (CEPRATE SC) and fluorescence-activated cell sorting: implications for gene transduction and/or transplantation.

The purification of early hematopoietic progenitor cells for autologous transplantation is based on two rationales: (1) elimination of clonogenic tumor cells, and/or (2) gene transfer into indefinitely self-replicating hematopoietic stem cells. Primitive CD34+ stem cells can be separated from more mature stem cells, or probably from clonogenic tumor cells, by differences in HLA-DR surface antigen expression. The objective of this study was to establish a large-scale technique for purification of CD34+, DR- progenitor cells from a large volume marrow harvest. In five different experiments, CD34+ cells were purified to between 76% and 91% by avidin-biotin immunoadsorption (CEPRATE SC) as a first step. This was followed by fluorescence-activated cell sorting to separate DR+ and DR- cells, which resulted in the generation of between 1.75 and 11.3 x 10(5) CD34+, DR- cells. The purity of DR- cells increased from between 0.5% and 4.3% in the immuno-adsorbed fraction up to 99% in the DR- sorted fraction. As shown in a single experiment, the purity of CD34+, DR- cells immediately after thawing increased from 0.01% to 94.3% while losing 99% of those early progenitor cells during the multistep purification procedure. We were able to physically separate one CD34+, DR- cell from up to 8000 nucleated cells in the prepurified cell suspension. One million highly purified CD34+, DR- progenitor cells is potentially an adequate cell dose for autologous transplantation equivalent to what is contained in an unselected and functioning marrow autograft.

Antigens, CD↗
PubMed1994

Karyotype and prognosis in non-Hodgkin lymphoma.

In malignant non-Hodgkin lymphomas (NHL), cytogenetic analysis may provide prognostic information including prediction of histologic evolution and responsiveness to therapy. In this study, we correlate clinical data and chromosomal aberrations in 70 adult patients with newly diagnosed NHL followed for a median of 20 months. Clonal aberrations were detected in 68/70 patients (97%). Besides t(2;5)(p23;q35), observed exclusively in three patients with anaplastic large cell lymphoma, Ki-1 positive, none of the characteristic aberrations observed was specific for a given histological subtype. Aberrations of chromosome 7 (n = 21) occurred in all histological subtypes together with aberrations of chromosome 3 and of the short arm of chromosome 17. They were clinically associated with a high serum lactate dehydrogenase level (LDH) and a trend to short survival. Anomalies of the long arm of chromosome 13 (n = 10) were found in patients with high grade B-cell lymphomas and bulky disease. In t(14;18)(q32;q21) bearing lymphomas (n = 27), distinct patterns of additional aberrations were observed in low grade and high grade lymphomas: trisomy 3 and trisomy 18 occurred concomitantly in high grade lymphomas (n = 6, p < 0.001) as well as aberrations of 1q, 5q, 6q and +der (18)(q21). In conclusion, cytogenetic analysis provides information about the complexity of genetic changes in NHL. These changes act not only as indicators of disease activity, but influence clinical outcome as demonstrated by their stringent correlation to the International Index and might reveal more general rules of tumor growth and spreading.

Adolescent↗
PubMed1993

Rapid induction of CD38 antigen on myeloid leukemia cells by all trans-retinoic acid.

The CD38 or T10 molecule is one of the least understood differentiation antigens. Virtually no information is available on the regulation and functions of CD38 antigen in hematopoietic cells. Using human promyelocytic leukemia cells, we demonstrate that all trans-retinoic acid is a potent and specific inducer of CD38 expression in myeloid lineage. At physiological doses, all trans-retinoic acid induces significant levels (8 to 10-fold) of CD38. Similarly, in patients with promyelocytic leukemia, a significant increase (3 to 6-fold) in CD38 expression was observed in vivo following single oral dose administration of all trans-retinoic acid. The induction of CD38 is a specific response of myeloid cells to retinoic acid and is not seen with other agents that induce differentiation. We believe that the induction of CD38 antigen is an early event in retinoid-regulated gene expression in normal and transformed myeloid cells.

ADP-ribosyl Cyclase↗
PubMed1993

Induction of differentiation in myeloid leukemia cell lines and acute promyelocytic leukemia cells by liposomal all-trans-retinoic acid.

All-trans-retinoic acid (ATRA) induces complete remissions in the majority of patients with acute promyelocytic leukemia. Despite continuous p.o. ATRA administration, many patients relapse after a short remission duration. In these patients, ATRA plasma concentrations were found to be very low, which was related to induction of retinoic acid-binding protein and increased drug catabolism by cytochrome P-450-mediated reactions. An i.v. ATRA formulation, which can be achieved by encapsulating ATRA into liposomes, may have the potential to overcome these unwanted effects. We investigated the ability of liposomal ATRA (L-ATRA) to induce differentiation of human myeloid leukemia cell lines (HL-60, KG-1, and THP-1). Cellular differentiation, as assessed by morphological criteria and by the expression of a mature myeloid cell surface antigen (CD11b on HL-60 and KG-1 cells), was induced by culture in the presence of L-ATRA. The ability of L-ATRA-treated HL-60 cells to reduce nitroblue tetrazolium demonstrated that they were functionally differentiated cells. Cell cycle analysis revealed significant growth inhibition of the cells after L-ATRA treatment. Following culture with L-ATRA, cells from five patients with acute promyelocytic leukemia were found to be morphologically and immunophenotypically mature myeloid cells. L-ATRA was as effective as free ATRA in inducing differentiation of the cell lines and of the cells from patients with acute promyelocytic leukemia. We conclude that L-ATRA effectively induces differentiation and may be a useful parenteral ATRA formulation for overcoming the pharmacological mechanisms that lead to "retinoid resistance."

Cell Cycle↗
PubMed1993

Flow cytometric detection of cytoplasmic antigens in acute leukemias: implications for lineage assignment.

This study aimed at optimizing the conditions for flow cytometric detection of myeloperoxidase (MPO), cytoplasmic CD3 (cCD3), and cytoplasmic CD22 (cCD22), which seem to be more reliable lineage-associated markers in acute leukemia than surface antigens. Fixation methods employing saponin as detergent resulted in accurate detection of MPO and cCD3, whereas cCD22 was detectable only after buffered-formaldehyde-acetone fixation. MPO was detected in 16/17 AML, but only in 1/6 ALL, the MPO positive ALL being also CD13 positive. MPO was detectable in 3/4 AML with T-lymphoid features; a case of myeloid antigen-positive T-ALL, however, was MPO-negative. cCD3 was expressed only in T-ALL, and five cases of lymphoid antigen-positive AML were cCD3-negative. We suggest that these flow cytometric assays are useful for the lineage assignment of poorly differentiated leukemias and contribute to the identification of truly biphenotypic acute leukemias.

Acute Disease↗
PubMed1993

Immunoglobulin gene rearrangement in plasma cell dyscrasias: detection of small clonal cell populations in peripheral blood and bone marrow.

The bone marrow (BM) and peripheral blood (PB) samples of 71 patients with plasma cell dyscrasias were analysed by the Southern blot technique for the presence of clonal immunoglobulin (Ig) gene rearrangements. 53% of BM samples examined were archival material such as air dried BM slides or frozen trephine biopsies. The results were related to bone marrow plasmacytosis as determined by cytology and flow cytometry, and other clinical parameters. Clonal Ig gene rearrangements were found in BM samples of 45 (83%) of 54 MM patients and in 3 of 6 patients with monoclonal gammopathy of unknown significance (MGUS). Clonal cell populations in the PB were detected in 11 (30%) of 37 examined MM patients, but in none of the patients with MGUS or solitary plasmacytoma of bone. PB involvement was associated with progressive disease. Circulating monoclonal cells were significantly associated with higher M-protein levels (p < 0.05). Thus, circulating clonal precursor cells are encountered more frequently in active MM.

Adult↗
PubMed1992

Subpopulations of normal peripheral blood and bone marrow cells express a functional multidrug resistant phenotype.

The multidrug-resistance gene, MDR1 is expressed in many normal tissues, but little is known about its expression in normal hematopoietic cells. Using the monoclonal antibody C219 and flow cytometric analysis, P-glycoprotein (P-gp) was found to be expressed in all peripheral blood (PB) subpopulations (CD4, CD8, CD14, CD19, CD56) except granulocytes. To specifically determine MDR1 gene expression, these PB subpopulations were isolated by fluorescence-activated cell sorting (FACS) and analyzed for MDR1 mRNA by polymerase chain reaction (PCR). All subsets were positive by PCR, but only minimal MDR1 mRNA was detected in monocytes and granulocytes. Significant efflux of Rhodamine-123 (Rh-123), a measure of P-gp function, was detected in CD4+, CD8+, CD14+, CD19+, and CD56+ cells but not in granulocytes. Next, PCR-analysis was performed on FACS-sorted bone marrow (BM) cells to assess MDR1 expression in different maturational stages. Precursors (CD34+), early and late myeloid cells (CD33+/CD34+, CD33+/CD34-) as well as lymphocytes of the B-cell lineage (CD19+/CD10+, CD19+/CD10-) expressed the MDR1 gene. BM monocytic cells (CD33++/CD34-) were negative, and a very weak signal was detected in erythroid cells (glycophorin A+). Significant Rh-123 efflux was found in CD34+, CD10+, CD33+, and CD33++ BM cells, but not in glycophorin A+ cells. We conclude that PB and BM lymphocytes, PB monocytes, BM progenitors, and immature myeloid cells, but not late BM monocytes, erythroid cells, and PB granulocytes, express MDR1 mRNA and a functional P-gp. These results have to be taken into account when MDR1 expression is determined in tumor samples containing normal blood cells.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1992

Proliferation of hematopoietic cells is accompanied by suppressed expression of heat shock protein 70.

In this study, we have examined the synthesis of heat shock protein (HSP70) in leukemia cells from acute myelogenous leukemia (AML) patients and in mononuclear cells from normal individuals, before and after growth factor stimulation. We have shown that the HSP70 protein was expressed in these cells in the absence of temperature elevation. Stimulation of proliferation of AML cells by the growth factors interleukin-3 and granulocyte-macrophage colony-stimulating factor and stimulation of normal lymphocytes by phytohemagglutinin (PHA) resulted in decreased synthesis of HSP70, suggesting that high levels of HSP70 are associated with cellular differentiation.

Bone Marrow↗
PubMed1992

Regulation of the expression of ICAM-1 on human monocytes and monocytic tumor cell lines.

In this study we investigated the mechanisms regulating expression of ICAM-1 that plays an important role for Ag presentation, on peripheral blood monocytes, and three myelomonocytic cell lines representing different stages of monocyte maturation. ICAM-1 expression on freshly isolated monocytes was low in all donors tested. After 16- to 20-h incubation, about a 20-fold increase was observed, whereas ICAM-1 expression on lymphocytes did not change. This marked increase in Ag expression was specific for ICAM-1, because expression of other monocyte Ag remained unchanged (alpha-chain of lymphocyte function-associated Ag-1) or decreased (ICAM-2, alpha-chains of CR3 and CR4, their common beta-chain and Fc gamma RI) during overnight incubation; only HLA-DR showed a slight increase. Enhanced expression of ICAM-1 was not caused by adherence; it was also not due to trace amounts of IFN-gamma possibly present under our culture conditions. Endotoxin contamination of the medium was excluded as a cause for the enhanced expression, and neither LPS nor its antagonist polymyxin B were able to alter ICAM-1 expression. Whole blood culture almost completely prevented up-regulation of ICAM-1 expression, thus apparently mimicking the in vivo situation. Culture of monocytes together with other PBMC had no significant influence on monocyte ICAM-1 levels. From 12 different cytokines tested for their ability to modulate "spontaneous" up-regulation of ICAM-1 in culture, only IFN-gamma was found to increase expression about twofold. This effect was also observed in whole blood culture. All other cytokines had no significant effect. In contrast, ICAM-1 expression on two of the three cell lines (KG1 and HL60) was inducible by TNF-alpha to a much higher degree than by IFN-gamma, whereas the results with U937 were comparable with those obtained for normal monocytes. The protein- and RNA-synthesis inhibitors cycloheximide and actinomycin D prevented expression of ICAM-1 in a dose-dependent fashion. In Northern blot experiments no difference in the amounts of mRNA was observed between freshly isolated and cultured monocytes, indicating that "spontaneous" induction of ICAM-1 is regulated at a posttranscriptional level. In contrast, stimulation with IFN-gamma caused a significant increase of detectable RNA comparable to that observed for Ag expression. These results disclose a very special regulation of ICAM-1 expression on monocytes which could be consistent with the view that ICAM-1 is actively down-regulated in vivo until the monocyte leaves the circulation.

Antigens, CD↗
PubMed1992

Flow cytometric determination of atypical antigen expression in acute leukemia for the study of minimal residual disease.

The aim of this study was to investigate to which extent acute leukemias could be monitored for residual disease by using atypical antigen combinations as leukemia-related markers. Atypical antigenic features were determined by double color flow cytometry and included coexpression of lymphoid and myeloid related antigens, unphysiological coexpression of immature and mature antigens, and lack of an antigen that is normally expressed during maturation. Atypical immunophenotypes were detected in 35 of 68 patients with AML (51.5%) and 15 of 24 patients with ALL (62.5%). When 12 patients with leukemia-associated markers were again analyzed at relapse, the relevant antigen combinations were retained in 11 of them. The sensitivity of this two color flow cytometric assay as determined in dilution experiments was 1 in 10(3) to 10(4) cells. Follow-up studies of bone marrow samples revealed that, after induction chemotherapy cells with leukemia-associated markers were detectable in several patients at a frequency of 0.5 to 4%, but only patients in whom the cells with atypical antigens never disappeared suffered from relapse. In contrast, patients who became negative for the atypical cells remained in complete remission (median remission duration after the first negative bone marrow assessment by flow cytometry 52 weeks, range 20-102). We conclude that atypical antigen combinations, which are present in a meaningful number of acute leukemias, are a valuable means of monitoring acute leukemia patients during follow-up. This flow cytometric approach can complement other strategies to get a more accurate definition of remission in acute leukemia.

Acute Disease↗
PubMed1992

Reversal of multidrug resistance by B859-35, a metabolite of B859-35, niguldipine, verapamil and nitrendipine.

It has been shown previously that verapamil and other calcium antagonists and calmodulin inhibitors can reverse multidrug resistance. We compared the potency of the dihydropyridine derivatives (4R)-3-[3-(4,4-diphenyl-1-piperadinyl)-propyl]-5-methyl-1,4-dihydr o-2,6- dimethyl-4-(3-nitrophenyl)-pyridine-3,5-dicarboxylate-hydrochloride (B859-35), a metabolite of B859-35, niguldipine and (R)-nitrendipine to that of (RS)-verapamil in reversing multidrug resistance. The accumulation of the fluorescent dye rhodamine 123, which is transported by the P-glycoprotein, was determined by a flow cytometer. Multidrug-resistant human HeLa KB-8-5 and Walker rat carcinoma cells were incubated in the presence and in absence of the drugs indicated above. We found that 0.1 microM B859-35 increases the accumulation of rhodamine 123 in multidrug-resistant KB-8-5 and Walker cells more effectively than 1 microM (RS)-verapamil. In sensitive KB-3-1 cells addition of the drugs had no significant influence on the accumulation of rhodamine 123. IN KB-8-5 cells, 10 nM Adriamycin caused a reduction of cell growth to 85% compared to untreated controls (= 100%). If 1 microM B859-35, B859-35 metabolite, niguldipine, verapamil or (R)-nitrendipine was added to 10 nM Adriamycin, growth reduction compared with untreated controls increased to 12%, 11%, 23%, 63%, and 82% respectively. The effect of 0.1 microM B859-35 was a reduction in proliferation to 38%, that of 0.1 microM verapamil to 72%. These data illustrate that B859-35, a compound with antitumor activity in several tumors, is at least ten times more potent than racemic verapamil in reversing multidrug resistance.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1992

Surface immunoglobulin density in the differential diagnosis of B-cell chronic lymphocytic leukemia and leukemic immunocytoma.

Using flow cytometry peripheral blood samples of 37 consecutive patients with B-cell chronic lymphocytic leukemia (B-CLL) and 17 consecutive patients with leukemic immunocytoma (IC) were studied in order to determine quantitative differences in the surface immunoglobulin (slg) density. In 8/37 (21.6%) cases of B-CLL and 1/17 (5.9%) cases of IC slg staining remained in the control level. Analysis of slg-positive cases demonstrated a close association between the amount of slg and diagnosis: per case the mean calculated fluorescence intensity for IC lymphocytes was 209.7 arbitrary linear intensity units (IU) (median: 156.4, standard error of the mean (SEM): 53.7) and for B-CLL lymphocytes 10.8 IU (median: 7.3, SEM: 1.1; p less than 0.0001). Altogether, 94.6% of all B-CLL patients and 76.5% of all IC patients were correctly classified when a cut-off point was fixed at a mean fluorescence intensity value of 20.0 IU. The percentage of leukemic cells as characterized by CD19 and HLA-DR reactivity was significantly lower in cases of IC (p less than 0.03 and p less than 0.01, respectively). In both entities disease progression occurred more frequently in advanced stages (II-IV) according to the Rai classification (p less than 0.01). In progressive disease rather than in stable disease circulating T lymphocytes were shown to express decreased amounts of surface CD3 antigen (p less than 0.02). We conclude that the quantitative assessment of surface antigens in addition to their qualitative characterization provides accurate information. In particular, the diagnostic discrimination between B-CLL and IC may be improved by determining the lymphocytes' slg amount.

Antigens, CD↗
PubMed1992

Detection of activity of P-glycoprotein in human tumour samples using rhodamine 123.

Based on the fluorescent properties of the dye rhodamine 123 (Rh123), which is transported by the membrane efflux pump P-glycoprotein (P-gp), we developed a functional flow cytometric assay for the detection of multidrug-resistant (MDR) cells. Using drug sensitive cell lines (KB-3-1) and MDR mutants (KB-8-5, KB-C1) experimental conditions were established that enabled demonstration of significant differences in Rh123 efflux and accumulation. Subsequently we investigated the applicability of this functional assay for the prediction of MDR in human peripheral blood and bone marrow samples. Using two-colour flow cytometry, the leukaemic blast cells of six patients suffering from acute myeloid leukaemia (AML) were analysed. In three cases the blast cells showed a rapid and marked Rh123 efflux. In the presence of MDR inhibitors these cells retained Rh123. To determine whether the efflux of Rh123 was associated with P-gp expression, the leukaemic cells were stained with the monoclonal antibody MRK-16. In addition extracted RNA was analysed by polymerase chain reaction to evaluate the expression of mdr 1 mRNA. In all three Rh123+ cases mdr 1 mRNA was detectable whereas only one AML case expressed P-gp. In comparing Rh123 with daunorubicin, which also allows the detection of MDR cells, accumulation studies proved Rh123 to be the more sensitive drug for flow cytometric MDR screening. Additionally, two-colour flow cytometry was much easier to perform with Rh123 than with daunorubicin. Our results indicate that flow cytometric measurement of Rh123 accumulation/efflux proves applicable to detect MDR cells in heterogenous clinical samples.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1991

Combined flow cytometric assessment of cell surface antigens and nuclear TdT for the detection of minimal residual disease in acute leukaemia.

To define more precisely the immunophenotype of lymphoid blast cells, a new flow cytometric technique for the simultaneous detection of surface antigens and nuclear terminal deoxynucleotidyl transferase (TdT) was established. After staining for the cell surface marker, mononuclear cells were treated with paraformaldehyde (1%) and methanol to permeabilize the cell membrane. Then the cells were stained by indirect immunofluorescence using a rabbit anti-human TdT antibody. Dilution experiments were performed to reveal the sensitivity of the described flow cytometric assay: 0.02% leukaemic cells could reliably be detected above background level among normal peripheral blood lymphocytes. It is concluded that the double-staining procedure described here is a sensitive tool that contributes to the detection of minimal residual disease in a substantial proportion of acute leukaemias.

Antigens, Neoplasm↗