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首页> 外文期刊>Cytometry: The Journal of the Society for Analytical Cytology >Competitive antibody binding to soluble CD16b antigen and CD16b antigen on neutrophils in whole blood by flow cytometry
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Competitive antibody binding to soluble CD16b antigen and CD16b antigen on neutrophils in whole blood by flow cytometry

机译：流式细胞术分析竞争抗体与全血中性粒细胞上可溶性CD16b抗原和CD16b抗原的结合

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Background: Shed receptors from the surface of white blood cells in in whole brood have been quantitated using the long and tedious enzyme-linked immunosorbent assay (ELISA) method. A simple rapid flow cytometric method of analysis for shed antigen in the presence of cell-bound antigen can be advantageous. Methods: Magnetic bead depletion of neutrophils in whole blood with CD16b antibody-conjugated beads as measured by flow cytometric analysis of the remaining cell suspension nas inhibited by the presence of soluble CD16b antigen in the brood plasma of normal donors. We describe a competitive binding assay between labeled and unlabeled CD16b antibody for receptors shed from the surface of formed bodies (cells) into solution. Also presented is a new method of obtaining the amount of soluble antigen in a sample. We determine the total unlabeled and labeled ligand concentration at which the fluorescence intensity of the labeled ligand matches the fluorescence intensity in a control run with only the labeled ligand. Results: Normal blood donors showed serum concentration levels of shed CD16b antigen in the range of 1-50 nM as determined by a flow cytometric competitive binding assay. These figures compared favorably with parallel determinations using magnetic bead depletion of targeted neutrophils for washed and unwashed whole blood samples to evaluate the concentration of shed CD16b antigen. Conclusions: The competitive antibody binding assay for shed and cell-bound CD16b antigen can be applied to similar GPI-linked antigens, for which purified antibody and fluorescent antibody against the same antigenic receptor are available. Cytometry 44:30-37, 2001. (C) 2001 Wiley-Liss, Inc.(dagger). [References: 23]
机译：背景：已经使用漫长而乏味的酶联免疫吸附测定（ELISA）方法对整个巢中白细胞表面的脱落受体进行了定量。在细胞结合抗原的存在下分析脱落抗原的简单快速流式细胞术方法可能是有利的。方法：通过流式细胞术分析剩余的细胞悬浮液的流式细胞仪分析，正常血液供体的血浆中存在可溶性CD16b抗原抑制，用CD16b抗体偶联的珠子测量全血中性粒细胞的磁珠消耗。我们描述了一种标记的和未标记的CD16b抗体之间竞争性结合测定，用于从形成的物体（细胞）表面脱落到溶液中的受体。还提出了一种获取样品中可溶性抗原量的新方法。我们确定总的未标记和标记的配体浓度，在该浓度下，标记的配体的荧光强度与仅使用标记的配体的对照运行中的荧光强度相匹配。结果：通过流式细胞术竞争结合测定法，正常献血者显示脱落的CD16b抗原的血清浓度水平在1-50 nM范围内。这些数字与平行磁法测定的嗜中性粒细胞的平行测定结果相吻合，平行测定法是针对洗过的和未洗过的全血样品评估脱落的CD16b抗原的浓度。结论：针对脱落和细胞结合的CD16b抗原的竞争性抗体结合试验可用于类似的GPI连接抗原，可获得针对相同抗原受体的纯化抗体和荧光抗体。 Cytometry 44：30-37，2001。（C）2001 Wiley-Liss，Inc。（匕首）。 [参考：23]

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