Different
leukemias express on their plasma membranes particular subsets of the 247 defined cluster of differentiation (
CD) antigens, which may resemble those of precursor cells along the lineages of differentiation to mature myeloid and lymphoid leukocytes. The extent of use of
CD antigen expression (immunophenotyping) for identification of
leukemias has been constrained by the technique used, flow cytometry, which commonly specifies only three
CD antigens in any one assay. Currently,
leukemias and
lymphomas are diagnosed using a combination of morphology, immunophenotype, cytochemistry, and karyotype. We have developed a rapid, simple procedure, which enables concurrent determination of 50 or more
CD antigens on leukocytes or
leukemia cells in a single analysis using a microarray of
antibodies. A
suspension of cells is applied to the array, and cells only bind to antibody dots for which they express the corresponding
CD antigen. For patients with significantly raised leukocyte counts, the resulting dot pattern then represents the immunophenotype of those cells. For patients at earlier stages of disease, the diagnosis depends on recognition of dot patterns distinct from the background of normal leukocytes. Distinctive and reproducible dot patterns have been obtained for normal peripheral blood leukocytes,
chronic lymphocytic leukemia (CLL),
hairy cell leukemia,
mantle cell lymphoma,
acute myeloid leukemia, and T-cell
acute lymphoblastic leukemia. The consensus pattern for
CD antigen expression found on CLL cells taken from 20 patients in descending order of cells bound was CD44,
HLA-DR, CD37, CD19, CD20, CD5, CD52, CD45RA, CD22, CD24, CD45, CD23, CD21, CD71, CD11c, and CD9. The
antigens that provided the best discrimination between CLL and normal peripheral blood leukocytes were CD19, CD20, CD21, CD22, CD23, CD24, CD25, and CD37. Results obtained for the expression of 48
CD antigens from the microarray compared well with flow cytometry. The microarray enables extensive immunophenotyping, and the intact cells captured on antibody dots can be further characterized using soluble, fluorescently labeled
antibodies.