Exocytotic exposure and retrieval of membrane antigens of chromaffin granules: quantitative evaluation of immunofluorescence on the surface of chromaffin cells.
Exocytotic exposure and retrieval of membrane antigens of chromaffin granules: quantitative evaluation of immunofluorescence on the surface of chromaffin cells.
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DOI：
10.1083/jcb.98.5.1817
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发表时间：
1984-05
期刊：
影响因子：
--
通讯作者：
Winkler H
中科院分区：
文献类型：
--
作者：
Patzak A;Böck G;Fischer-Colbrie R;Schauenstein K;Schmidt W;Lingg G;Winkler H
用从牛肾上腺髓质分离的嗜铬细胞研究了嗜铬颗粒膜抗原的胞吐暴露。使用抗糖蛋白 III 和多巴胺 β-羟化酶的抗血清，通过间接膜免疫荧光观察细胞表面的抗原。对于未刺激的细胞，仅在细胞表面观察到微弱的免疫荧光，而刺激的细胞（使用卡巴胆碱或 Ba2+）表现出更强的反应。在所有情况下，染色均表现为点和斑。为了定量证明这些观察结果，我们使用荧光激活细胞分选仪分析了免疫染色的细胞。刺激后，细胞群的平均荧光强度增强。这种增加与儿茶酚胺分泌的程度相关。受刺激细胞的荧光强度在很大范围内变化，表明各个细胞对促分泌剂的反应各不相同。当刺激后的细胞在 37°C 下孵育长达 45 分钟时，观察到膜免疫荧光的减少接近未刺激的对照细胞。显然，已经融入质膜的嗜铬颗粒膜是通过特定且相对快速的过程回收的。这种从细胞表面回收抗原的过程被叠氮化钠阻断，但不受秋水仙碱、细胞松弛素 B 和三氟拉嗪的影响。本文建立的定量方法对于进一步研究分泌组织中的胞吐-内吞循环动力学应该是有用的。
The exocytotic exposure of antigens of chromaffin granule membranes was studied with chromaffin cells isolated from bovine adrenal medulla. Antigens on the cell surface were visualized by indirect membrane immunofluorescence employing antisera against glycoprotein III and dopamine beta-hydroxylase. With unstimulated cells, only weak immunofluorescence on the cell surface was observed, whereas stimulated cells (with carbachol or Ba2+) exhibited much stronger reactions. In all cases the staining appeared as dots and patches. To quantitatively prove these observations, we analyzed the immunostained cells using a fluorescence-activated cell sorter. After stimulation, the average fluorescence intensity of the cell population was enhanced. This increase correlated with the degree of catecholamine secretion. The fluorescence intensity of stimulated cells varied over a broad range indicating that individual cells reacted variably to the secretagogues. When stimulated cells were incubated at 37 degrees C for up to 45 min after stimulation, a decrease of membrane immunofluorescence approaching that of unstimulated control cells was observed. Apparently, the membranes of chromaffin granules, which had been incorporated into the plasma membrane, were retrieved by a specific and relatively fast process. This retrieval of the antigen from the cell surface was blocked by sodium azide, but not influenced by colchicine, cytochalasin B, and trifluoperazine. The quantitative methods established in this paper should prove useful for further study of the kinetics of the exo-endocytotic cycle in secretory tissues.
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影响因子：
7.3
作者：
DOUGLAS, WW
通讯作者：
DOUGLAS, WW
影响因子：
4.7
作者：
HUBER, E;KONIG, P;WINKLER, H
通讯作者：
WINKLER, H
DOI：
10.1073/pnas.74.11.5031
发表时间：
1977-01-01
影响因子：
11.1
作者：
BOURGUIGNON, LYW;SINGER, SJ
通讯作者：
SINGER, SJ
影响因子：
64.8
作者：
LINGG, G;FISCHERCOLBRIE, R;WINKLER, H
通讯作者：
WINKLER, H
影响因子：
7.8
作者：
POSNER, BI;VERMA, AK;BERGERON, JJM
通讯作者：
BERGERON, JJM