Negative immunomagnetic selection of T cells from peripheral blood of presentation AML specimens.
To date, studies on T cells in acute myeloid leukemia (AML) have been limited to flow cytometric analysis of whole peripheral blood mononuclear cell (PBMC) specimens or functional work looking at the impact of AML myeloblasts on normal or remission T cells. This lack of information on T cells at the time of presentation with disease is due in part to the difficulty in isolating sufficiently pure T cells from these specimens for further study. Negative immunomagnetic selection has been the method of choice for isolating immune cells for functional studies due to concerns that binding antibodies to the cell surface may induce cellular activation, block ligand-receptor interactions or result in immune clearance. In order specifically to study T cells in presentation AML specimens, we set out to develop a method of isolating highly pure CD4 and CD8 T cells by negative selection from the peripheral blood (PB) of newly diagnosed AML patients. This technique, unlike T cell selection from PB from normal individuals or from patients with chronic lymphocytic leukaemia, was extremely problematic due to properties of the leukaemic myeloblasts. A successful method was eventually optimized requiring the use of a custom antibody cocktail consisting of CD33, CD34, CD123, CD11c and CD36, to deplete myeloblasts.
Le Dieu
Rifca
R
Institute of Cancer, Centre for Medical Oncology, Barts and The London School of Medicine, Queen Mary University of London, Charterhouse Square, London EC1M 6BQ UK.
Taussig
David
D
Lister
T Andrew
TA
Gribben
John G
JG
D000906:Antibodies / Q000276:immunology; D015703:Antigens, CD / Q000276:immunology; D018952:Antigens, CD34 / Q000276:immunology; D015214:Antigens, Differentiation, Myelomonocytic / Q000276:immunology; D039521:CD11c Antigen / Q000276:immunology; D018955:CD36 Antigens / Q000276:immunology; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology*; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology*; D005434:Flow Cytometry; D042381:Granulocyte Precursor Cells / Q000276:immunology; D006801:Humans; D018189:Immunomagnetic Separation / Q000379:methods*; D053650:Interleukin-3 Receptor alpha Subunit / Q000276:immunology; D015470:Leukemia, Myeloid, Acute / Q000097:blood / Q000276:immunology*; D063268:Sialic Acid Binding Ig-like Lectin 3
D000906:Antibodies; D015703:Antigens, CD; D018952:Antigens, CD34; D015214:Antigens, Differentiation, Myelomonocytic; D039521:CD11c Antigen; C571163:CD33 protein, human; D018955:CD36 Antigens; C115022:IL3RA protein, human; D053650:Interleukin-3 Receptor alpha Subunit; D063268:Sialic Acid Binding Ig-like Lectin 3