Biomedical subjects

R Morilla

Publications and source records attributed to R Morilla.

At least 37 records · Page 2Linked to original sources

PubMed1996

Differential expression of CD3 and CD7 in T-cell malignancies: a quantitative study by flow cytometry.

Most T-cell antigens are expressed on normal and neoplastic T lymphocytes and for this reason it is not easy to distinguish between the immunophenotype of normal and malignant T cells. We have addressed this problem by comparing the levels of expression of CD3 and CD7 on T lymphocytes from 18 healthy donors with those of 61 cases of T-cell leukaemia using quantitative flow cytometry with a method that converts fluorescence intensity into number of antigen molecules per cell. Normal T lymphocytes expressed 124 +/- 25 CD3 and 20 +/- 3 x 10(3) CD7 molecules per cell. The mean CD3 values were significantly lower in all types of T-cell leukaemia than in normal T cells (P < 0.05), with the exception of Sezary syndrome. The lowest CD3 values were found in T-lymphoblastic leukaemia (T-ALL), 30 +/- 21 x 10(3), and adult T-cell leukaemia/lymphoma (ATLL), 38 +/- 31 x 10(3), followed by T-prolymphocytic leukaemia (T-PLL), 92 +/- 47 x 10(3), and granular lymphocyte leukaemia (GLL), 95 +/- 21 x 10(3). In contrast, the number of CD7 molecules was significantly higher in T-All, 35 +/- 7 x 10(3) (P < 0.01), and T-PLL, 29 +/- 12 x 10(3), than the normal controls (P < 0.01), whereas ATLL and GLL showed a low CD7 expression, 13 +/- 3 and 12 +/- 3 x 10(3), respectively. Our results show that the quantitative analysis of CD3 and CD7 and their combined evaluation may enable a distinction between normal and leukaemic T cells and could facilitate the monitoring of minimal residual disease. This study has also defined the T prolymphocyte as a cell of intermediate maturity between thymic derived and peripheral T lymphocytes.

Antigens, CD7↗

PubMed1996

The ultrastructure of mantle cell lymphoma and other B-cell disorders with translocation t(11;14)(q13;q32).

We conducted an ultrastructural study in 22 cases of B-lymphoproliferative disorders in leukaemic phase bearing the t(11;14) translocation. The features of peripheral blood leukaemic cells in nine cases of mantle cell lymphoma (MCL) were compared to those diagnosed as B-prolymphocytic leukaemia (B-PLL) (five cases), splenic lymphoma with villous lymphocytes (SLVL) (four cases), lymphoplasmocytic lymphoma (LPL) (one case), chronic lymphocytic leukaemia with > 10% prolymphocytes (CLL/ PL) (one case) and unclassified B-non Hodgkin's lymphoma (B-NHL) (two cases). The ultrastructural characteristics were also compared to those present in B-NHL without t(11;14), including cases of follicular centre lymphoma (FCL). This study shows that MCL has distinct ultrastructural features including a cleaved or indented nucleus with an even heterochromatin distribution, an absent or inconspicuous nucleolus, low N/C ratio, abundant mitochondria, a well developed Golgi zone, profiles of endoplasmic reticulum and centrioles. This pattern clearly differs from that found in FCL cells. The nuclear pattern of MCL cells also differed from the cells in the other disorders with t(11;14), but shared an organelle-rich cytoplasm, and features which were not apparent in cases without t(11;14). The cytoplasmic changes observed in cells bearing t(11;14) suggest increased cellular activity which may relate to the chromosome translocation and the resulting over-expression of bcl-1.

B-Lymphocytes↗

PubMed1996

Expression of TIA-1 and TIA-2 in T cell malignancies and T cell lymphocytosis.

OBJECTIVE: To investigate the reactivity with TIA-1 and TIA-2, two monoclonal antibodies that recognise, respectively, granular structures in T lymphocytes and the T cell receptor chain in cells from a variety of T cell disorders. METHODS: Cytoplasmic staining with TIA-1 and TIA-2 was carried out by the immunoalkaline phosphatase anti-alkaline phosphatase technique in 67 cases with a T cell disorder: 31 large granular lymphocyte (LGL) leukaemia, nine T-prolymphocytic leukaemia (T-PLL), five Sezary syndrome, four peripheral T cell lymphoma (PTCL), 13 T cell lymphocytosis, and five T-acute lymphoblastic leukaemia (T-ALL). All had over 75% abnormal T cells which were CD2+, CD3+, CD5+, CD7+, and negative with B cell markers. RESULTS: TIA-1 was positive in 77% cases of LGL leukaemia and half of the PTCL and T-ALL, whereas it was negative in all Sezary syndrome and most T-PLL (8/9) and reactive T-lymphocytosis (10/13). In LGL leukaemia, TIA-1 was positive irrespective of the membrane phenotype, whether CD8+, CD4- or CD4+, CD8-, and was more often positive in cases where cells were CD16+, CD56+, or CD57+. TIA-2 was positive in 60% of cases encompassing all diagnostic types of T cell disorder. There was no correlation between TIA-2 expression and that of other T cell markers, activation antigens, and natural killer markers. CONCLUSIONS: The pattern of TIA-1 expression in T cell malignancies may help in the differential diagnosis among LGL leukaemia (high expression), T cell lymphocytosis and other T cell diseases (low expression). As TIA-2 is expressed in over 95% mature T lymphocytes and thymic cells, its assessment may be useful to demonstrate aberrant phenotypes which can be exploited for detecting minimal residual disease.

Antibodies, Monoclonal↗

PubMed1996

Differential expression of T cell antigens in normal peripheral blood lymphocytes: a quantitative analysis by flow cytometry.

AIMS: To obtain reference values of the level of expression of T cell antigens on normal lymphocyte subsets in order to disclose differences which could reflect their function or maturation stages, or both. METHODS: Peripheral blood from 15 healthy donors was processed by flow cytometry with triple colour analysis. For each sample phycoerythrin (PE) conjugated CD2, CD4, CD5, CD8, and CD56 monoclonal antibodies were combined with Cy5-R-phycoerythrin (TC) conjugated CD3 and fluorescein isothiocyanate (FITC) conjugated CD7; CD2- and CD7-PE were also combined with CD3-TC and CD4-FITC. Standard microbeads with different capacities to bind mouse immunoglobulins were used to convert the mean fluorescence intensity (MFI) values of the lymphocyte subsets identified by multiparametric flow cytometry into the number of antigen molecules per cell, measured as antibody binding capacity (ABC). RESULTS: CD4+ (helper/inducer) T cells exhibit a higher CD3 antigen expression compared with CD8+ (suppressor/ cytotoxic) T lymphocytes. Within the CD4+ T cells, the CD4+CD7- subset expressed a lower level of CD3 compared with CD4+CD7+ and CD8+CD7+ cells, and higher CD2 and CD5 expression than the main CD3+CD7+ subset. Major differences in antigen expression were also detected between CD3+ T cells and CD3-CD56+ natural killer (NK) cells: NK cells exhibited higher levels of CD7 and CD56 and lower levels of CD2 and CD5 than T cells. Significantly lower CD5 expression was also detected in the small CD5+ B lymphocyte subset compared with T cells. CONCLUSIONS: Quantitative flow cytometry with triple colour analysis may be used to detect antigen modulations in disease states and to increase the accuracy of diagnosis by comparison with findings in normal counterparts.

Adult↗

PubMed1996

Zeta chain and CD28 are poorly expressed on T lymphocytes from chronic lymphocytic leukemia.

We have analyzed the expression of the zeta chain of the T cell receptor/CD3 complex and the co-stimulatory molecule CD28 by dual colour immunofluorescence on T lymphocytes from patients with B cell chronic lymphocytic leukemia (CLL). Zeta chain was significantly reduced on CD3-positive lymphocytes from 33 patients compared with normal controls (P<0.0001). The values were lower in stages B and C than in stage A. In five patients tested in partial remission the values were normal. CD28, investigated in CD3, CD4 and CD8 positive T cells from 18 CLL patients appeared to be reduced in the three subsets but more marked in CD8-positive lymphocytes. The loss of zeta chain and CD28 in a proportion of circulating T lymphocytes from CLL may underlie some of the known functional abnormalities of these cells and the immunodeficiency associated with the disease.

CD28 Antigens↗

PubMed1996

Differential expression of B29 (CD79b) and mb-1 (CD79a) proteins in acute lymphoblastic leukaemia.

CD79 is a heterodimeric molecule comprising two polypeptide chains, B29 (CD79b) and mb-1 (CD79a). It is physically linked in the surface of B cells to membrane immunoglobulin, forming the B cell antigen receptor complex. Expression of the mb-1 (CD79a) chain has been studied in leukaemias and shown to be present in most B lineage acute lymphoblastic leukaemias (ALL). In contrast, little is known about the expression of B29 (CD79b) in this condition. Two monoclonal antibodies (MoAb) were used in this study by immunocytochemistry and flow cytometry: HM57, against an intracellular epitope of the mb-1(CD79a) chain, and SN8, reacting with an extracellular epitope of B29 (CD79b). Our aim was to investigate the expression of B29 (CD79b) in the various immunological subtypes of B lineage ALL and compare its cytoplasmic and membrane expression. Seventy-nine cases were studied, including 13 chronic myeloid leukaemia in B lymphoid blast crisis (CML-BC) and 66 ALL, subclassified as early B (two), common (28), pre-B (23), mature (five) and biphenotypic with B lymphoid commitment (eight). Most cases expressed mb-1 (CD79a) in the cytoplasm. B29 (CD79b) was expressed in the cytoplasm in 65% (15/23) of pre-B-ALL and in 14% (4/28) common-ALL but it was detected in the cell membrane in only three cases of mature B-ALL, being negative in all other B lineage subtypes ALL. Three of the biphenotypic leukaemias coexpressed cytoplasmic B29 (CD79b) and mu-chain. This was also seen in two cases of CML-BC, while four cases expressed only cytoplasmic B29 (CD79b) without mu-chain. Our results suggest that during B cell differentiation, B29 (CD79b) is expressed later than mb-1 (CD79a) in the cytoplasm and parallels the cytoplasmic expression of mu-chain. B29 (CD79b) is present in the membrane at a later stage compared to its cytoplasmic expression and found in mature B blasts (B-ALL) that express membrane Ig as it is in normal and leukaemic B lymphocytes.

Antibodies, Monoclonal↗

PubMed1996

Concurrent activation of MYC and BCL2 in B cell non-Hodgkin lymphoma cell lines by translocation of both oncogenes to the same immunoglobulin heavy chain locus.

Concurrent activation of BCL2 and MYC usually occurs in B cell non-Hodgkin lymphoma (B-NHL) by translocation of both oncogenes to both immunoglobulin heavy chain (IGH) alleles: this abrogates immunoglobulin synthesis. We have studied three B-NHL cell lines (DoHH2, VAL and ROS 50) and show that concurrent activation of BCL2 and MYC may follow translocation of both oncogenes to the same IGH allele. Conventional cytogenetics of DoHH2 suggested the presence of a t(14;18)(q32;q21) translocation. However, fluorescent in situ hybridization (FISH) studies using whole chromosome paints, alpha satellite probes and flow-sorted chromosomes as probes revealed an unexpected complexity of rearrangements involving chromosomes 8, 14 and 18, namely t(8;14;18)(q24;q32;q21). DNA blot and previous PCR analysis confirmed the juxtaposition of BCL2 major breakpoint region (mbr) with IGJH6, but also demonstrated a rearrangement within the first exon of MYC. The centromeric (5') MYC rearranged fragment comigrated with the BCL2-JH6 rearranged fragment in BamHI, EcoRI and Bg/II restriction digests. The der(8)t(8;14;18) therefore comprised 5' MYC (exon I)-Sgamma4-JH6-BCL2 mbr. Similar rearrangements were observed in both ROS 50 and VAL cell lines which contained two and three copies of the der(8)t(8;14;18) respectively. Quantitative flow cytometry for BCL2 and MYC expression showed abundant expression of both proteins in all three lines. These data indicate the der(14)t(14;18)(q32;q21) may itself be the target for any second translocation. The presence of the intact BCL2-JH fusion gene on the der(8)t(8;14;18) allowed concurrent activation of both BCL2 and MYC with no loss of immunoglobulin expression.

Aged↗

PubMed1996

Cytogenetic findings in acute biphenotypic leukaemia.

We have studied the immunological and cytogenetic features of 26 patients with acute leukaemia classified as biphenotypic according to a scoring system based on the number and lineage specificity of antigens expressed on the blast cells. The series included 19 adults (age >15 years) and seven children. The cases were distributed in four immunophenotypic groups: (1)coexpression of myeloid and B antigens, 18 cases (69 percent);(2)myeloid and T cell antigens, six (23 percent); (3) one case with trilineage differentiation; and (4) one case with coexpression of both B and T cell antigens. Cytogenetic analysis revealed a normal karyotype in four cases (15 percent) and abnormal clones in 22 (85 percent). Eight patients had the Philadelphia (Ph) translocation, t(9;22)(q34;q11), (31 percent), three cases had structural aberrations of 6q and two had 11q23 rearrangements, one with t(11;19) and a second with t(4;11); the other eight cases had different alterations including t(9;12)(q1;q1), t(8;21)(q22;q22), t(2;7) (p1?3;q3?4), t(7;12)(q11;p11), hyperdiploidy and other structural abnormalities. The chromosomal rearrangements in children were characterised by abnormalities of 11q23 in two cases and the Ph translocation in three. Our data indicate that biphenotypic features are common in cases presenting with t(9;22) as the eight cases included here represent 47 percent of all cases of Ph+ve acute leukaemia studied in our Institution. Biphenotypic acute leukaemias comprise a heterogeneous group of leukaemias involving pluripotent stem cells. Cytogenetic studies are essential in characterising these cases as they will disclose several poor prognosis chromosome aberrations of which the Ph chromosome is the most frequent.

Adolescent↗

PubMed1996

Expression of the immunoglobulin-associated protein B29 in B cell disorders with the monoclonal antibody SN8 (CD79b).

We studied the expression of the immunoglobulin-associated membrane protein B29 in 499 cases of chronic B cell diseases using the monoclonal antibody SN8 (CD79b). SN8 was positive in 5% (17/330) of chronic lymphocytic leukemia (CLL) and 100% (15/15) of B prolymphocytic leukemia. The expression of B29 in other B cell disorders was, as a rule, significantly higher than in CLL. Two thirds of non-Hodgkin's lymphomas in leukemic phase were SN8 positive, including lymphoplasmacytic (45%), follicular (83%), mantle cell (92%) and splenic lymphoma with villous lymphocytes (74%) while only 25% of hairy cell leukemias were SN8 positive. Within CLL, 2.3% of typical cases were SN8+ while 16% of cases with atypical morphology and an increased number of prolymphocytes were SN8+. Our results suggest a useful role for SN8 in the immunophenotypic differentiation of B cell disorders as a marker for non-CLL diseases. The analysis of B29 expression may throw light into the structure of the B cell antigen receptor in B cell malignancies while the distinctive reactivity profile of SN8 has direct applications to diagnosis.

Antibodies, Monoclonal↗

PubMed1995

Quantitative flow cytometry can distinguish between normal and leukaemic B-cell precursors.

The immunological detection of minimal residual disease in B-lineage acute lymphoblastic leukaemia (ALL) has been hampered by the fact that the leukaemic cells represent the malignant counterparts of normal haemopoietic precursors expressing terminal deoxynucleotidyl transferase (TdT), CD10 and CD19. We have used quantitative double-labelling flow cytometry with standard fluorescent beads to convert the mean fluorescence to the number of antigen molecules per cell. The number of TdT, CD10 and CD19 molecules per cell was determined in normal B-cell precursors from 22 healthy donors and eight regenerating marrows from patients with various malignancies and in 20 cases of B-lineage ALL. In normal bone marrow we characterized two different B-cell populations: TdT+/CD10+/CD19+ and TdT-/CD10+/CD19+. We demonstrated a major difference in the level of expression of TdT, CD10 and CD19 between normal bone marrow and B-lineage ALL blasts. Normal TdT+ precursors have significantly higher number of TdT (> 100 x 10(3)) and lower number of CD10 (< 50 x 10(3)) and CD19 (< 10 x 10(3)) molecules per cell than B-lineage ALL blasts (< 100, > 50, > 10 x 10(3) molecules per cell respectively); these differences were statistically highly significant. Furthermore, regenerating marrows had a significantly higher percentage of B-cell precursors than healthy donors. This increase was at the expense of the TdT-/CD10+/CD19+ population which, in the context of B-lineage ALL, could be wrongly interpreted as evidence of relapse if TdT is not included in the analysis. Therefore the quantitative analysis of TdT combined with CD10 and CD19 may allow a clear distinction between normal precursors and minimal residual leukaemia in B-lineage ALL and avoid the pitfall of misinterpreting regenerating B-cells as evidence of relapse.

Adolescent↗

PubMed1995

Differential TdT expression in acute leukemia by flow cytometry: a quantitative study.

Terminal deoxynucleotidyl transferase (TdT) has long been considered a diagnostic marker for acute lymphoblastic leukemia. Reports of TdT-positive cells in acute myeloid leukemia have lately questioned its diagnostic value. TDT has been detected mainly by microscopy methods: immunofluorescence and immunocytochemistry. The aim of this study was to reevaluate the diagnostic importance of TdT in acute leukemia by using flow cytometry with a method that allows quantitative analysis. Fifty-eight cases of acute leukemia were studied and TdT expression was quantified using calibrated fluorescent beads. The highest TdT values were found in B lineage acute lymphoblastic leukemia (ALL) while acute myeloid leukemia (AML) had the lowest values, even in cases with a high percentage of TdT-positive cells. Biphenotypic leukemia had intermediate values between B-lineage and T-lineage acute leukemia. The difference between these groups was statistically significant (P < 0.0001). The TdT assay by flow cytometry was more precise than immunocytochemistry because it recognizes quantitative differences between ALL and AML. It is also valuable in better defining the maturation stages in pre-B ALL and T-ALL. We conclude that quantitative flow cytometry of TdT re-establishes the diagnostic value of this enzyme and has potential applications for the study of minimal residual disease.

Acute Disease↗

PubMed1995

[Phenotypical diagnosis of Japanese chronic lymphocytic leukemia--an international cooperative study based on the French-American-British classification of mature B-cell leukemias].

To better understand the accuracy in the diagnosis of chronic lymphocytic leukemia (CLL) and characterize the clinical features of CLL in Japan, where the disease is extremely rare, an international cooperative study was conducted by hematologists between Japan and UK. Blood and bone marrow films from 36 patients with a possible-diagnosis of CLL were referred to two laboratories of Nagasaki University Hospital (Nagasaki) and Royal Marsden Hospital (London). According to the FAB criteria, typical CLL 16 and CLL/PL 2, a subtype of atypical CLL, were completely accordant in diagnosis. However, phenotypical diagnosis of CLL mixed, the other of atypical CLL, and intermediate lymphocytic leukemia (ILL) in leukemic phase often gave inconsistent results. Especially, 8 cases of atypical CLL designated as likely CLL were equivocal between CLL and ILL, suggesting clinical feature more close to typical CLL than ILL. This indicates the presence of a relatively high incidence of atypical CLL in Japan which dose not exactly fit with the FAB Criteria. Finally, we would like to emphasize that an international cooperative study allows improvement of accuracy in diagnosis and better understanding of the disease entity of lymphoid malignancies, having on ethnically different morbidity.

Aged↗

PubMed1995

Comparison of Cobe Spectra and Haemonetics MCS-3P cell separators for peripheral blood stem cell harvesting.

A prospective study was undertaken to compare the total nucleated cell (TNC), mononuclear cell (MNC), CD34+ cell, and CFU-GM yields of two different cell separators. A Haemonetics MCS-3P and a Cobe Spectra machine were used to leukapherese 10 patients with malignant diseases on 4 consecutive days after mobilization with G-CSF. All patients were harvested twice on each machine for a fixed period of time. The blood volume processed (10.47 vs 3.79 l, P < 0.001), MNC yield (2.66 vs 0.90 x 10(8)/kg; P < 0.001), MNC yield rate (1.66 vs 0.55 x 10(6)/kg/min; P < 0.001), MNC purity (81 vs 42%; P < 0.001), CFU-GM yield (18.1 vs 5.5 x 10(4)/kg; P = 0.001), and CFU-GM yield rate (11.27 vs 3.42 x 10(2)/kg/min; P = 0.001) were significantly higher with the Spectra. The CD34+ cell yields and yield rates were comparable. Although CFU-GM and MNC yields per unit blood volume processed were comparable for both machines, there was a trend for higher CD34+ yields per unit volume processed with MCS-3P. We conclude that Spectra is faster than MCS-3P with more blood processed per unit time resulting in higher cell yields, but yields per unit volume processed are comparable for both machines. The choice of machine for a given patient depends upon convenience, venous access and the time available.

Adult↗

PubMed1994

B-cell non-Hodgkin's lymphoma cell line (Karpas 1106) with complex translocation involving 18q21.3 but lacking BCL2 rearrangement and expression.

A B-cell non-Hodgkin's lymphoma (B-NHL) cell line (Karpas 1106) with an unusual three-way translocation involving 18q21.3 has been derived from a patient with mediastinal lymphoblastic B-NHL. Although conventional cytogenetics showed a derivative 18q-identical to that seen in cases with t(14;18)(q32.3;q21.3), no translocations of either chromosome 14 could be detected. Instead fluorescent in situ hybridization analysis using a chromosome-18 paint showed that the segment 18q21.3-18qter had become sandwiched on a derivative chromosome X between segments Xqter-c-Xq28 and 13q12-qter, with the centrometric site of 18q21.3 subband juxtaposed to the X sequences. Pulsed-field DNA blots failed to detect rearrangement of the BCL2 gene. Conventional DNA blots using a variety of restriction digests and both 5' and 3' BCL2 and FVT 1 probes also failed to detect rearrangement in Karpas 1106. A rearranged fragment seen only in HindIII digests with 5' BLC2 probes may represent a local microalteration, which is either a mutation or small deletion involving the HindIII site as seen in other cases of B-NHL. Neither BCL2 RNA nor BCL2 protein expression were detected. These and other data suggest that genes at 18q21.3, other than BCL2 and FVT1, may be targets for translocation in certain subgroups of B-NHL.

Adult↗

PubMed1994

The immunophenotype of splenic lymphoma with villous lymphocytes and its relevance to the differential diagnosis with other B-cell disorders.

Splenic lymphoma with villous lymphocytes (SLVL) is a low-grade disorder that regularly presents with peripheral blood involvement. We describe the immunophenotype of the circulating cells from 100 SLVL patients whose disease has been characterized on clinical, morphologic, and histologic grounds. Cells from all cases expressed B-cell antigens (CD19 and CD37) and/or HLA-Dr and showed light chain restriction (kappa/lambda: 1.5/1) with moderate to strong intensity of membrane Ig staining. Cells from most cases (> 80%) were CD24+, FMC7+, and expressed strongly membrane CD22. The monoclonal antibodies CD10, CD23, and CD38 were positive in one-third of the cases; CD11c in 47%; and CD25 in 25% of cases. A minority of cases (< 20%) were positive with HC2, B-ly-7, and CD5. However, none of the 19 CD5+ cases had the phenotype characteristic of chronic lymphocytic leukemia (CD5+, CD23+, FMC7-, weak surface Ig and membrane CD22). None of the 17 CD25+ cases had the immunophenotype typical of hairy cell leukemia (CD25+, CD11c+, HC2+, B-ly-7+). HC2 and B-ly-7 were the most useful reagents to distinguish SLVL from hairy cell leukemia. Our findings demonstrate that SLVL has a distinct immunologic profile and that monoclonal antibodies are important for the differential diagnosis between this disease and other B-lymphoproliferative disorders with which SLVL can be confused.

Adult↗

PubMed1994

Trisomy 12 in B-cell chronic lymphocytic leukaemia: assessment of lineage restriction by simultaneous analysis of immunophenotype and genotype in interphase cells by fluorescence in situ hybridization.

We have studied the lineage restriction of trisomy 12 in six patients with B-cell chronic lymphocytic leukaemia (CLL) by simultaneous analysis of immunophenotype and fluorescence in situ hybridization (FISH) signals in single interphase cells. Fresh uncultured cells from each patient were immunophenotyped by the alkaline phosphatase anti-alkaline phosphatase method (APAAP) using monoclonal or polyclonal antibodies and hybridized with a chromosome 12 specific alpha-satellite DNA probe. In all cases trisomy 12 was restricted to the clonal B-cells, kappa positive or lambda positive, whereas T-cells (CD3 positive) and non clonal B-cells had only two chromosome 12 signals. Within the clonal B-cell population a large proportion of cells were disomic for chromosome 12, whilst trisomic cells ranged from 21% to 37%. The absence of trisomy 12 in T-cells and the mosaicism demonstrated in the clonal B-cells suggests that this abnormality is a secondary event during the leukaemic transformation of CLL and develops in an already established neoplastic B-cell population.

Aged↗

PubMed1994

Demonstration of cytoplasmic and nuclear antigens in acute leukaemia using flow cytometry.

AIMS: To detect cytoplasmic and nuclear antigens using flow cytometry in acute leukaemia and to use this technique for double marker combinations. METHODS: Cytoplasmic staining was carried out in samples from 40 cases of acute leukaemia with monoclonal antibodies against the myeloid antigen CD13, the lymphoid antigens CD3, CD22, mu chain and the enzymes terminal deoxynucleotidyl transferase (TdT) and myeloperoxidase (MPO). The cells were fixed with paraformaldehyde and permeabilised with Tween 20 and Becton Dickinson's FACS lysing solution. Flow cytometry results were compared in the same cases with immunocytochemistry results using the alkaline phosphatase anti-alkaline phosphatase method. RESULTS: The gentle permeabilisation induced by this method permitted preservation of the membrane antigens and the size and morphology of the cells. The results using flow cytometry were comparable with those obtained using immunocytochemistry, with nearly complete concordance in most cases. CONCLUSIONS: This technique is simple, rapid, sensitive and reproducible and it is suitable for double staining procedures, such as nuclear and cytoplasmic, nuclear and membrane, or cytoplasmic and membrane. It therefore provides a powerful tool for extending the use of immunophenotyping for the diagnosis and follow up of acute leukaemia. It could also be used for the investigation of minimal residual disease.

Acute Disease↗

PubMed1994

Growth of primary human acute lymphoblastic and myeloblastic leukemia in SCID mice.

The in vivo growth of 8 human primary acute lymphoblastic (ALL) and 17 primary acute myeloblastic (AML) cell populations was investigated in severe combined immunodeficient (SCID) mice. Bone marrow (BM) or peripheral blood (PB) samples, either fresh or cryopreserved, were implanted i.v. into irradiated SCID mice. The cells from 5/8 patients with ALL resulted in engraftment with systemic proliferation and dissemination leading to morbidity and mortality of the animals within 12-18 weeks from implantation. In contrast, none of the 17 AML samples resulted in sustained engraftment. Four of the 5 engrafted ALL populations have been successfully passaged into fresh recipients and phenotypic and karyotypic characteristics remain unaltered. The data presented indicate that the SCID mouse may be a useful model for studying the pathogenesis of ALL and may also enable the development and investigation of new therapies for this disease.

Adult↗