Novel method for immunofluorescence staining of mammalian eggs using non-contact alternating-current electric-field mixing of microdroplets.
Novel method for immunofluorescence staining of mammalian eggs using non-contact alternating-current electric-field mixing of microdroplets.
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DOI：
10.1038/srep15371
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发表时间：
2015-10-19
影响因子：
4.6
通讯作者：
Terada Y
中科院分区：
文献类型：
--
作者：
Shirasawa H;Kumagai J;Sato E;Kabashima K;Kumazawa Y;Sato W;Miura H;Nakamura R;Nanjo H;Minamiya Y;Akagami Y;Terada Y
最近，开发了一种用于非催化混合微滴的新技术。在该方法中，使用交流（AC）电场来促进微滴内的抗原-抗体反应。以前，该技术仅应用于扁平结构的组织学检查，例如手术标本。在这项研究中，我们首次将这种技术应用于三维结构的免疫荧光染色，特别是哺乳动物的卵子。我们将抗微管抗体从1：1,000稀释到1：16,000，并比较了不同稀释度的色度和褪色程度。此外，我们改变交流电场混合的频率从5 Hz到46 Hz，并评估对微管染色的影响。在AC电场混合仅5分钟后，微管被更强烈地染色，即使当一抗的浓度比常规方法低10倍。交流电场混合也减轻微管褪色。在所有测试频率下，AC电场混合导致比对照组更强的微管染色。频率之间的微管染色没有明显差异。这些结果表明，新方法可以减少抗体消耗，缩短免疫荧光染色时间。
Recently, a new technique was developed for non-catalytically mixing microdroplets. In this method, an alternating-current (AC) electric field is used to promote the antigen–antibody reaction within the microdroplet. Previously, this technique has only been applied to histological examinations of flat structures, such as surgical specimens. In this study, we applied this technique for the first time to immunofluorescence staining of three-dimensional structures, specifically, mammalian eggs. We diluted an antibody against microtubules from 1:1,000 to 1:16,000, and compared the chromatic degree and extent of fading across dilutions. In addition, we varied the frequency of AC electric-field mixing from 5 Hz to 46 Hz and evaluated the effect on microtubule staining. Microtubules were more strongly stained after AC electric-field mixing for only 5 minutes, even when the concentration of primary antibody was 10 times lower than in conventional methods. AC electric-field mixing also alleviated microtubule fading. At all frequencies tested, AC electric-field mixing resulted in stronger microtubule staining than in controls. There was no clear difference in a microtubule staining between frequencies. These results suggest that the novel method could reduce antibody consumption and shorten immunofluorescence staining time.