DiOC₂(3) 绿色膜电位荧光探针

产品关键词：

DiOC2(3)；DiOC6(3)；壬基吖啶橙NAO；JC-10 线粒体膜电位探针；MitoTracker® Green FM 绿色线粒体荧光探针；罗丹明123 Rhodamine 123；CAS NO：905-96-4；

产品信息

【温馨提示】：见我司懋康生物（maokangbio）提供的线粒体荧光探针产品专题。

产品描述

DiOC2(3)，英文全名3,3'-Diethyloxacarbocyanine Iodide，CAS NO. 905-96-4，是一种膜电位探针，可通过流式细胞仪检测根据发射波长荧光信号比来分析细菌活力。DiOC2(3)常用来测定细菌膜电位，也能用于研究某些活细胞比如大鼠胚胎成纤维细胞、猴肾细胞、中国仓鼠肺成纤维细胞和小鼠3T3成纤维细胞。

DiOC2(3)用来检测细菌膜电位的工作原理在于：DiOC2(3)在所有细菌细胞内发绿色荧光，由于更高的膜电位引起染料分子发生自聚作用，使得染料的荧光往红色发射波长处迁移，红色荧光强度增高，此时红色/绿色荧光信号比高。质子离子载体比如CCCP，通过去除质子梯度来破坏线粒体膜电位，从而导致红色荧光强度降低，此时红色/绿色荧光信号比降低。DiOC2(3)的最大激发和发射波长约482nm和497nm。经DiOC2(3)标记的细胞用流式细胞仪分析（用488nm激发，分别用适合FITC（绿色）和Texas Red（红色）的发射滤片来检测）。

产品特性

1）   CAS NO.：905-96-4

2）   化学名：3-ethyl-2-[3-(3-ethyl-2(3H)-benzoxazolylidene)-1-propenyl]-benzoxazolium iodide

3）   同义名：3,3'-Diethyloxacarbocyanine Iodide 3,3′-二乙基氧杂羰花青碘; DiOC2(3) Iodide; DOC Iodide;

4）   分子式：C21H21IN2O2

5）   分子量：460.31

6）   纯度：>95.0% (HPLC)

7）   Ex/Em：482/497nm（in methanol）

8）   外观：红色粉末或结晶

9）   溶解性：溶于DMSO，DMF，无水乙醇，微溶于水

10）化学结构式：

保存与运输方法

保存：-20℃避光干燥保存，至少一年有效。

运输：室温运输。

注意事项

1）   荧光染料均存在淬灭问题，请尽量注意避光，以减缓荧光淬灭。

2）   为了您的安全和健康，请穿实验服并戴一次性手套操作。

使用方法

1.染色液制备

1）储存液制备：用 DMSO或无水乙醇配置浓度1～5 mM的储存液。例如，取10mg DiOC2(3)（Mw：460.31 g/mol）溶于4.35ml 无水DMSO中，充分溶解，即得到5mM的储存液。【注意】未使用的储存液分装储存在-20℃，避免反复冻融，且要避光保存。

2）工作液制备：用合适的缓冲液（如：无血清培养基，HBSS或PBS）稀释储存液，调整到合适的工作浓度。【注意】：工作液最终浓度需要根据不同细胞系和实验体系来优化。

2.染色方法

对于细菌染色，最好取对数生长期的健康细菌，用无菌PBS或其他适当平衡盐缓冲液稀释到~1 x 106cells/ml。也可直接从细菌培养液中取适量的细菌不经清洗进行稀释。细菌染色用DiOC2(3)的工作浓度可为30μM，室温染色15~30min。

3.染色示例

Fig 1. Detection of membrane potential in mycobacteria. Red/green ratios were calculated using population mean fluorescence intensities for mycobacteria, incubated with 3 μM DiOC2 for 30 min in either the presence or absence of 25 μM CCCP.

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参考文献

[1]Chen Z et al. Hypoionic Shock Facilitates Aminoglycoside Killing of Both Nutrient Shift- and Starvation-Induced Bacterial Persister Cells by Rapidly Enhancing Aminoglycoside Uptake. Front Microbiol. 2019 Sep 6;10:2028. PMID: 31551965

A flow cytometry-based assay was applied to measure the PMF by using the fluorescence probe 3,3′-Diethyloxacarbocyanine Iodide [DiOC2(3); purchased from MaoKang Biotechnology, Inc., Shanghai, China] according to the manufacturer’s instruction. Briefly, E. coli persisters, with or without CCCP pretreatment as described above, were diluted into PBS to a cell density of 106 cells/mL and incubated with DiOC2(3) (at a final concentration of 30 μM) at room temperature for 30 min. Cells were subjected to flow cytometric analysis on FACSymphonyTMA5 (BD Biosciences) with an excitation at 488 nm and emission at both red and green channels.

[2]Zhao Y et al. Rapid Freezing Enables Aminoglycosides To Eradicate Bacterial Persisters via Enhancing Mechanosensitive Channel MscL-Mediated Antibiotic Uptake. mBio. 2020 Feb 11;11(1). pii: e03239-19. doi: 10.1128/mBio.03239-19. PMID: 32047133

【3】A flow cytometry-based assay was applied to measure the proton motive force by using the fluorescence probe 3,3′-diethyloxacarbocyanine iodide [DiOC2(3)] (purchased from MaoKang Biotechnology, Inc., Shanghai, China) according to the manufacturer’s instructions. Briefly, E. coli exponential-phase cells, with or without CCCP pretreatment for 1 h, were diluted into PBS to a cell density of 106 cells/ml and incubated with DiOC2(3) (at a final concentration of 30 μM) at room temperature for 15 min. Cells were subjected to flow cytometric analysis on a FACSymphony A5 system (BD Biosciences) with excitation at 488 nm and emission in both the red (630-nm) channel and green (515-nm) channel. Cells treated by rapid freezing for 10 s with liquid nitrogen were also analyzed.

[4]Participation of the inositol 1,4,5-trisphosphate-gated calcium channel in the zona pellucida- and progesterone-induced acrosome reaction and calcium influx in human spermatozoa.

Hoechst 33258 and polyvinylpyrrolidone-40 (PVP-40) were obtained from Shanghai Maokang Biotechnology Co., Ltd. (Shanghai, China). The spermatozoa were washed and incubated with Hoechst 33258 (4 μg ml−1) for 10 min at a proportion of 9:1 to achieve a final concentration of 0.4 μg ml−1 and then centrifuged with 2% (w/v) PVP-40 in PBS. The supernatant was then removed and resuspended in PBS, smeared on clean slides, and fixed for 30 min in 95% (v/v) ethanol after air drying.

 — —Written/Edited by V. Shallan【版权归MKBio懋康所有】

上海懋康生物科技有限公司是一家涉足于生命科学和生物技术领域研究的试剂、仪器和实验室消耗品与实验服务工作，主要从事细胞生物学、植物学、分子生物学、免疫学、生物化学、蛋白组学。生物制药与诊断试剂研发生产等领域。 本公司秉承“以人为本，以诚为信、合同守信”的经营理念。坚持"品质保障"的原则为广大客户提供优质产品。