Combined immunophenotyping and DNA in situ hybridization to study lineage involvement in patients with myelodysplastic syndromes

Abstract Clonality of myeloid and lymphoid cell fractions obtained from peripheral blood (PB) or bone marrow (BM) of five patients with a myelodysplastic syndrome (MDS), was studied by combined immunophenotypic analysis and DNA in situ hybridization. This novel technique enables quantitative and direct analysis of cytogenetic alterations in nondividing cells of distinct cell lineages. Four patients with a trisomy 8 and one patient with a translocation (1;7) were studied. For cell lineage determination, antibodies specific for progenitor cells (CD34), myeloid cells (CD15), monocytes (63D3), T cells (CD3), and B cells (CD19,20,22) were used. In one patient with a trisomy 8, BM cells were available and the erythroid lineage could be studied. For detection of cytogenetic aberrations, we used chromosome- specific repetitive DNA probes. In three patients, all nonlymphoid cells carried the cytogenetic abnormality; in two patients, mosaicism within these lineages was suggested by the relative low numbers (35% to 55%) of aberrant cells. None of the T or B cells of the five patients carried the chromosomal aberrations. We conclude that combined immunophenotyping and in situ hybridization is a feasible technique to study lineage involvement. Our data suggest that the chromosomal aberrations studied in MDS are restricted to the myeloid lineages.

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Combined immunophenotyping and DNA in situ hybridization to study lineage involvement in patients with myelodysplastic syndromes

Blood ◽

10.1182/blood.v79.7.1823.bloodjournal7971823 ◽

1992 ◽

Vol 79 (7) ◽

pp. 1823-1828 ◽

Cited By ~ 1

Author(s):

RE Kibbelaar ◽

H van Kamp ◽

EJ Dreef ◽

G de Groot-Swings ◽

JC Kluin-Nelemans ◽

...

Keyword(s):

In Situ Hybridization ◽

B Cells ◽

Chromosomal Aberrations ◽

Cell Lineage ◽

Direct Analysis ◽

Trisomy 8 ◽

Cytogenetic Aberrations ◽

Distinct Cell ◽

Cell Fractions

Clonality of myeloid and lymphoid cell fractions obtained from peripheral blood (PB) or bone marrow (BM) of five patients with a myelodysplastic syndrome (MDS), was studied by combined immunophenotypic analysis and DNA in situ hybridization. This novel technique enables quantitative and direct analysis of cytogenetic alterations in nondividing cells of distinct cell lineages. Four patients with a trisomy 8 and one patient with a translocation (1;7) were studied. For cell lineage determination, antibodies specific for progenitor cells (CD34), myeloid cells (CD15), monocytes (63D3), T cells (CD3), and B cells (CD19,20,22) were used. In one patient with a trisomy 8, BM cells were available and the erythroid lineage could be studied. For detection of cytogenetic aberrations, we used chromosome- specific repetitive DNA probes. In three patients, all nonlymphoid cells carried the cytogenetic abnormality; in two patients, mosaicism within these lineages was suggested by the relative low numbers (35% to 55%) of aberrant cells. None of the T or B cells of the five patients carried the chromosomal aberrations. We conclude that combined immunophenotyping and in situ hybridization is a feasible technique to study lineage involvement. Our data suggest that the chromosomal aberrations studied in MDS are restricted to the myeloid lineages.

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Comparison of the fetal and adult functional B cell repertoires by analysis of VH gene family expression.

Journal of Experimental Medicine ◽

10.1084/jem.168.2.589 ◽

1988 ◽

Vol 168 (2) ◽

pp. 589-603 ◽

Cited By ~ 71

Author(s):

H D Jeong ◽

J M Teale

Keyword(s):

Gene Expression ◽

In Situ Hybridization ◽

B Cells ◽

Gene Family ◽

B Cell ◽

Cell Lineage ◽

Cell Repertoire ◽

B Cell Repertoire ◽

Vh Gene

The functional B cell repertoire in BALB/c mice was assessed at various stages in ontogeny. This was done by analyzing VH gene family expression using the sensitive technique of in situ hybridization. The B cell repertoire was probed with the mitogen, LPS, and the antigen DNP. DNP was chosen because B cells responsive to this hapten appear very early in ontogeny. The APCs that developed after stimulation with LPS or DNP were analyzed for VH gene expression by in situ hybridization of individual cells using radiolabeled VH gene family probes. The results indicated that VH gene expression in fetal B cells after stimulation was distinct from adult B cells in that there was a biased expression of D proximal families. The results indicated that this bias was associated with developmental age and not a given differentiation stage in the B cell lineage. In addition, stimulation of fetal B cells with DNP resulted in a large increase in expression of member(s) of VH 36-60, suggesting that the early appearance of DNP-responsive B cells is not strictly correlated with preferential rearrangement of D proximal families, VH 7183 and VH Q52. However, the results suggested that a large proportion of pre-B cells that preferentially rearrange D proximal families early in ontogeny become part of the functional developing repertoire.

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In situ hybridization on May-Grunwald Giemsa-stained bone marrow and blood smears of patients with hematologic disorders allows detection of cell-lineage-specific cytogenetic abnormalities

Blood ◽

10.1182/blood.v82.3.884.bloodjournal823884 ◽

1993 ◽

Vol 82 (3) ◽

pp. 884-888 ◽

Cited By ~ 1

Author(s):

K van Lom ◽

A Hagemeijer ◽

EM Smit ◽

B Lowenberg

Keyword(s):

Bone Marrow ◽

In Situ Hybridization ◽

Cell Morphology ◽

Chromosomal Abnormalities ◽

Cell Lineage ◽

Hematologic Malignancies ◽

Cytogenetic Aberrations ◽

Cytotoxic Treatment ◽

Before And After

Bone marrow and blood from patients with acute myeloid leukemia and myelodysplastic syndrome were studied by simultaneous analysis of cell morphology and karyotype. A combined technique of May-Grunwald Giemsa (MGG) for cell morphology and fluorescence in situ hybridization (FISH) with chromosome-specific DNA probes for detection of cytogenetic aberrations allowed us to investigate cell-lineage-specific chromosomal abnormalities. We introduced video recordings to examine large numbers of cells. Briefly, evaluation was first performed on MGG slides, during which cell position and morphology were recorded on an S-VHS recorder. Subsequently, the same slides were used for FISH. This resulted in the identification of MGG-stained cells on the video screen and, at the same time, the interpretation of FISH signals in the fluorescence microscope. Specimens of bone marrow or blood samples from four patients with different hematologic malignancies were studied. One of these patients was studied before and after cytotoxic treatment. The gain or loss of chromosomes could be detected easily and morphologically assigned to the blasts in all patients and to a variable proportion of the myelomonocytic lineage in two patients, but not to the lymphocytes. Thus, this method provides new possibilities for investigating the clonality of hematologic malignancies.

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Cell lineage involvement in four patients with myelodysplastic syndrome and t(1;7) or trisomy 8 studied by simultaneous immunophenotyping and fluorescence in situ hybridization

Cancer Genetics and Cytogenetics ◽

10.1016/0165-4608(93)90180-t ◽

1993 ◽

Vol 70 (2) ◽

pp. 120-124 ◽

Cited By ~ 27

Author(s):

Stella J. Nylund ◽

Walter Verbeek ◽

Marcelo L. Larramendy ◽

Tapani Ruutu ◽

Kristiina Heinonen ◽

...

Keyword(s):

In Situ Hybridization ◽

Myelodysplastic Syndrome ◽

Fluorescence In Situ Hybridization ◽

Cell Lineage ◽

Trisomy 8

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In situ hybridization on May-Grunwald Giemsa-stained bone marrow and blood smears of patients with hematologic disorders allows detection of cell-lineage-specific cytogenetic abnormalities

Blood ◽

10.1182/blood.v82.3.884.884 ◽

1993 ◽

Vol 82 (3) ◽

pp. 884-888 ◽

Cited By ~ 43

Author(s):

K van Lom ◽

A Hagemeijer ◽

EM Smit ◽

B Lowenberg

Keyword(s):

Bone Marrow ◽

In Situ Hybridization ◽

Cell Morphology ◽

Chromosomal Abnormalities ◽

Cell Lineage ◽

Hematologic Malignancies ◽

Cytogenetic Aberrations ◽

Cytotoxic Treatment ◽

Before And After

Abstract Bone marrow and blood from patients with acute myeloid leukemia and myelodysplastic syndrome were studied by simultaneous analysis of cell morphology and karyotype. A combined technique of May-Grunwald Giemsa (MGG) for cell morphology and fluorescence in situ hybridization (FISH) with chromosome-specific DNA probes for detection of cytogenetic aberrations allowed us to investigate cell-lineage-specific chromosomal abnormalities. We introduced video recordings to examine large numbers of cells. Briefly, evaluation was first performed on MGG slides, during which cell position and morphology were recorded on an S-VHS recorder. Subsequently, the same slides were used for FISH. This resulted in the identification of MGG-stained cells on the video screen and, at the same time, the interpretation of FISH signals in the fluorescence microscope. Specimens of bone marrow or blood samples from four patients with different hematologic malignancies were studied. One of these patients was studied before and after cytotoxic treatment. The gain or loss of chromosomes could be detected easily and morphologically assigned to the blasts in all patients and to a variable proportion of the myelomonocytic lineage in two patients, but not to the lymphocytes. Thus, this method provides new possibilities for investigating the clonality of hematologic malignancies.

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