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这是Cytometry A杂志2013年三月份的新文章，PE（藻红蛋白）是流式抗体上很常用的一种荧光标记，亮度高，应用广。本文作者发现PE标记的抗体，在小鼠固有层浆细胞（取自小鼠小肠粘膜）胞内染色时出现明显的非特异性染色，而FITC和APC标记的抗体则不会。这种非特异性染色与小鼠种类、实验方法、抗体来源种类都无关，仅跟PE标记有关。另外，体外活化的小鼠B细胞亦可发现这种非特异性染色。
非特异性染色
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＝＝＝＝原文摘要＝＝＝＝
Cytometry A. 2013 Mar 5. doi: 10.1002/cyto.a.22276. [Epub ahead of print]
R-phycoerythrin-conjugated antibodies are inappropriate for intracellular staining of murine plasma cells.
Kim MS , Kim TS .
Source
Division of Life Sciences, School of Life Sciences and Biotechnology, Korea University, Seoul 136-701, Republic of Korea.
Abstract
Phycoerythrin (PE) is a type of phycobiliproteins found in cyanobacteria and red algae. PE-conjugated antibodies are broadly used for flow cytometry and immunofluorescence microscopy. Because nonspecific binding of antibodies results in decreased analytic accuracy, numerous efforts have been made to unveil cases and mechanisms of nonspecific bindings. However, nonspecific binding of specific cell types by a fluorescent dye-conjugated form of antibody has been rarely reported. In the present study, we discovered that PE-conjugated antibodies, but not FITC- or APC-antibodies, selectively stained lamina propria plasma cells (LP-PCs) from the murine small intestine after membrane permeabilization. We demonstrated that LP-PC-selective staining with PE-antibodies was not due to interactions of antibody-epitope or antibody-Fc receptor. This unexpected staining by PE-antibody was not dependent on the mouse strain of LP-PCs, experimental methods, or origin species of the antibody, but dependent on PE itself. This phenomenon was also observed in plasma cells isolated from bone marrow, spleen, and mesenteric lymph nodes. Furthermore, in vitro activated B cells and in vivo generated LP-PCs were also selectively stained by PE-conjugated antibodies. Taken together, these results show that PE-conjugated antibodies are inappropriate for intracellular staining of murine plasma cells. © 2013 International Society for Advancement of Cytometry.
PMID:23463627
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发表于 2013-3-19 14:24:11
发表于 2013-6-21 08:02:33
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