Belov L, de la Vega O, dos Remedios C G, Mulligan S P, Christopherson R I
Department of Biochemistry, Institute for Biomedical Research, University of Sydney, NSW 2006, Australia.
Cancer Res. 2001 Jun 1;61(11):4483-9.
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.
不同的白血病在其质膜上表达247种已定义的分化簇（CD）抗原的特定亚群，这些亚群可能类似于沿分化谱系至成熟髓样和淋巴样白细胞的前体细胞的抗原亚群。用于白血病鉴定的CD抗原表达（免疫表型分析）的应用程度一直受到所使用技术——流式细胞术的限制，该技术在任何一次检测中通常只能指定三种CD抗原。目前，白血病和淋巴瘤的诊断采用形态学、免疫表型、细胞化学和核型分析相结合的方法。我们开发了一种快速、简单的程序，能够在一次分析中使用抗体微阵列同时测定白细胞或白血病细胞上的50种或更多种CD抗原。将细胞悬液应用于微阵列，细胞只会与它们表达相应CD抗原的抗体点结合。对于白细胞计数显著升高的患者，所得的点模式代表这些细胞的免疫表型。对于疾病早期阶段的患者，诊断取决于识别与正常白细胞背景不同的点模式。已获得正常外周血白细胞、慢性淋巴细胞白血病（CLL）、毛细胞白血病、套细胞淋巴瘤、急性髓细胞白血病和T细胞急性淋巴细胞白血病的独特且可重复的点模式。从20名患者采集的CLL细胞上发现的按结合细胞降序排列的CD抗原表达的共识模式为CD44、HLA-DR、CD37、CD19、CD20、CD5、CD52、CD45RA、CD22、CD24、CD45、CD23、CD21、CD71、CD11c和CD9。在CLL和正常外周血白细胞之间提供最佳区分的抗原是CD19、CD20、CD21、CD22、CD23、CD24、CD25和CD37。从微阵列获得的48种CD抗原表达结果与流式细胞术结果相当吻合。该微阵列能够进行广泛的免疫表型分析，并且捕获在抗体点上的完整细胞可以使用可溶性荧光标记抗体进行进一步表征。
