CD200R surface expression as a marker of murine basophil activation.
CD200R surface expression as a marker of murine basophil activation.
复制标题
DOI：
10.1111/j.1365-2222.2008.03154.x
复制
发表时间：
2009-03
期刊：
影响因子：
--
通讯作者：
Mitre E
中科院分区：
文献类型：
--
作者：
Torrero MN;Larson D;Hübner MP;Mitre E
嗜碱性粒细胞在变态反应性疾病和蠕虫感染的免疫应答中发挥着重要作用。研究嗜碱性粒细胞的主要障碍之一是缺乏一种简单快速的测定小鼠嗜碱性粒细胞活化的方法。本研究的目的是开发一种测定小鼠嗜碱性粒细胞活化的方法。用嗜碱性粒细胞的阳性和阴性标志物的各种组合对小鼠血细胞进行染色-分选，然后通过细胞离心涂片的May-Grüunwald染色评估嗜碱性粒细胞纯度。一旦确定了嗜碱性粒细胞染色的流式细胞术策略，在用抗IgE、离子霉素或N-甲酰基MetLeuPhe（fMLP）刺激全血后，通过多色流式细胞术评估CD 200 R的嗜碱性粒细胞表面表达。通过细胞内IL-4的伴随细胞染色来评估嗜碱性粒细胞活化的确认。为了测试流式细胞术嗜碱性粒细胞CD 200 R测量评估抗原特异性IgE介导的嗜碱性粒细胞活化的能力，在从对照小鼠、蠕虫感染的小鼠和对卵清蛋白致敏的小鼠获得的嗜碱性粒细胞上测量了响应于单独培养基、蠕虫抗原或卵清蛋白的体外刺激的表面CD 200 R表达。使用抗IgE-FITC作为阳性标记物，抗CD 4-PERCP和抗B220-PERCP的组合作为阴性标记物，导致嗜碱性粒细胞群体分离良好。用抗CD 200 R-PE进行的额外染色证明：（1）嗜碱性粒细胞CD 200 R表达响应于抗IgE、离子霉素和fMLP而增加，（2）大多数CD 200 R阳性嗜碱性粒细胞也对IL-4呈阳性染色，以及（3）在蠕虫病和变态反应的鼠模型中嗜碱性粒细胞的抗原特异性活化后，CD 200 R表达增加。我们开发了一种多色流式细胞术测定，利用CD 200 R作为活化标志物来测量鼠嗜碱性粒细胞活化。该检测方法简单快速，取血、体外刺激和流式细胞仪分析大约需要半天时间。
Basophils are increasingly recognized as playing important roles in the immune responses of allergic diseases and helminth infections. One of the main obstacles to studying basophils has been the lack of a simple and rapid assay to measure basophil activation in mice. The purpose of this study was to develop an assay to measure murine basophil activation. Mouse blood cells were stained with various combinations of positive and negative markers for basophils – sorted and then assessed for basophil purity by May-Grüunwald staining of cytospins. Once a flow cytometric strategy for staining basophils was determined, basophil surface expression of CD200R was assessed by multi-colour flow cytometry after stimulation of whole blood with anti-IgE, ionomycin or N-formyl MetLeuPhe (fMLP). Confirmation of basophil activation was assessed by concomitant staining of cells for intracellular IL-4. To test the ability of flow cytometric basophil CD200R measurements to assess for antigen-specific IgE-mediated activation of basophils, surface CD200R expression in response to in vitro stimulation with media alone, helminth antigen or ovalbumin was measured on basophils obtained from control mice, mice infected with helminths and mice sensitized to ovalbumin. Using anti-IgE-FITC as a positive marker and a combination of anti-CD4-PERCP and anti-B220-PERCP as negative markers resulted in a well-separated basophil population. Additional staining with anti-CD200R-PE demonstrated that (1) basophil CD200R expression increases in response to anti-IgE, ionomycin and fMLP, (2) most CD200R-positive basophils also stain positively for IL-4 and (3) CD200R expression increases after antigen-specific activation of basophils in murine models of helminth disease and allergy. We developed a multi-colour flow cytometry assay that measures murine basophil activation by utilizing CD200R as an activation marker. This assay is straightforward and rapid, taking approximately half a day for obtaining blood, in vitro stimulation and flow cytometric analysis.
登录
查看更多内容
影响因子：
4.4
作者：
Kojima, Toshiyuki;Obata, Kazushige;Karasuyama, Hajime
通讯作者：
Karasuyama, Hajime
影响因子：
4.4
作者：
Mitre, E;Taylor, RT;Nutman, TB
通讯作者：
Nutman, TB
影响因子：
4.4
作者：
Gorczynski, R;Chen, ZQ;Marsden, PA
通讯作者：
Marsden, PA
DOI：
10.1159/000088886
发表时间：
2006-01-01
期刊：
PARASITES AND ALLERGY
影响因子：
--
作者：
Mitre, Edward;Nutman, Thomas B.
通讯作者：
Nutman, Thomas B.
影响因子：
4.4
作者：
Cherwinski, HM;Murphy, CA;Phillips, JH
通讯作者：
Phillips, JH