A method for cloning T‐lymphocytic precursors in agar

Löwenberg, Bob; Zeeuw, H. M. C. de

doi:10.1002/ajh.2830060106

Cited by 33 publications

(4 citation statements)

References 18 publications

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“…This inhibition of myeloma growth by agar is not unique, we have experienced a similar inhibitory effect when culturing normal human 'T' cell colonies. Furthermore, the method developed by Lowenberg & De Zeeuw (1979) for 'T' cell growth utilizes a liquid overlay. Although the growth of myeloma colonies was maximal in the absence of agar, colony counting was difficult due to the tendency of cells to grow to confluence.…”

Section: Methodsmentioning

confidence: 99%

A simple method for culturing myeloma cells from human bone marrow aspirates and peripheral blood in vitro

et al. 1988

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A double layer agar technique has been developed to grow myeloma colonies (MY-CFUc) from human bone marrow aspirates and peripheral blood. Heavily irradiated HL60 cells (5 x 10(5)/plate) are added to an agar underlay in growth medium containing 0.5% agar. Mononuclear cells from the test bone marrow or blood are overlayered in either 0.2 ml HL60-conditioned medium (HL60-CM) or in 0.5 ml growth medium containing 0.23% agar, and the cultures are incubated at 37 degrees C in an atmosphere of 5% CO2, 10% O2 and 85% N2. Colonies (greater than 50 cells) form between 2 and 3 weeks. Using this method 60/68 samples of bone marrow and 7/12 samples of blood from 54 patients have produced colonies in soft agar and in liquid on an agar underlay. The cells which form these colonies are of two distinct sizes, the larger cells being plasmacytoid and the smaller lymphoid. The two cell types are usually, but not always, present in separate colonies. Both plasmacytoid and lymphoid cells carry the isotype of the respective patient's myeloma protein and the plasma cell marker (HAN PC1). This technique has enabled us to culture myeloma cells from patients with as few as 2% plasma cells in the bone marrow but it does not permit the growth of normal B, T or granulocyte-macrophage colonies (GM-CFUc). The drug sensitivity of myeloma cells (MY-CFUc) compared with normal haemopoietic cells (GM-CFUc) can be measured using dose-response curves in individual patients. Furthermore, this method can detect resistant subpopulations within a given myeloma sample.

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Section: Methodsmentioning

confidence: 99%

A simple method for culturing myeloma cells from human bone marrow aspirates and peripheral blood in vitro

et al. 1988

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“…Therefore, the T-lymphocytes in these cell preparations included both viable and nonviable cells that had been lysed specifically by the action of complement and fixed by the monoclonal antibody preparation used in the reaction mixture. These T-lymphocytes were cultured using a modification of Lowenberg and De Zeeuw's method [14]. Mononuclear cells (1 X 105) were cultured in 0.4 ml of McCoy's, containing 10% heat-inactivated FCS and 10 p1 of phytohemagglutinin-M (PHA; Wellcome Reagents Ltd., Kent, England), and layered over a 1 ml underlayer of McCoy's, containing 10% heat-inactivated FCS and 0.5% agar in 30 mm plastic Petri dishes (NUNC Laboratories, Copenhagen, Denmark).…”

Section: T-colony Formationmentioning

confidence: 99%

Depletion of T‐lymphocyte subsets using monoclonal antibody and complement: Effect on T‐colony formation

McCluskey

Kelly

Morris

et al. 1984

The International Journal of Cell Cloning

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Using monoclonal antibody and fresh rabbit serum, mononuclear cells from 11 healthy individuals were depleted of either helper or suppressor T-lymphocytes by complement-mediated lysis. The effect on T-lymphocyte colony formation was studied to determine the nature of the T-colony-forming cell. While depletion of either subset of T-lymphocytes significantly reduced T-colony formation compared to controls (P less than 0.01), colony formation did not differ significantly between helper or suppressor T-cell-depleted cells. Thus T-colony formation appears to be a property of both helper and suppressor T-lymphocytes, and both subsets are necessary for optimal colony formation.

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“…Cell Culture T lymphocytes were cultured using a modifica tion of the technique described by Lowenberg and De Zeeuw [1979]. In brief, 2.5 x 104 to 2.0 x 10s mono nuclear cells were cultured in 0.4 ml of McCoy's 5A medium containing 10% heat-inactivated fetal calf serum (Gibco) and 10 p\ of phytohaemagglutinin-M (PHA), over a l-ml underlayer of McCoy's 5A medi um containing 10% heat-inactivated fetal calf serum and 0.5% agar.…”

Section: Cell Separationmentioning

confidence: 99%

Quantitative and Qualitative Deficiency of T-Lymphocyte Colony Formation in T-Cell Lymphoma-Leukaemia

Kelly¹,

Morris

McCluskey

1983

Acta Haematol

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T-lymphocyte colony formation in a patient with T-cell lymphoma-leukaemia was found to be defective in three ways. Colony numbers in patient cultures were reduced below those of 20 normal donors. Colony size in patient cultures was uniformly small. Colony formation by patient cells was inhibited in co-cultures of patient and normal cells. These observations indicate that the leukaemic T cells in this rare entity have a defective T-cell function.

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A method for cloning T‐lymphocytic precursors in agar

Cited by 33 publications

References 18 publications

A simple method for culturing myeloma cells from human bone marrow aspirates and peripheral blood in vitro

A simple method for culturing myeloma cells from human bone marrow aspirates and peripheral blood in vitro

Depletion of T‐lymphocyte subsets using monoclonal antibody and complement: Effect on T‐colony formation

Quantitative and Qualitative Deficiency of T-Lymphocyte Colony Formation in T-Cell Lymphoma-Leukaemia

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