DiOC₂(3) 綠色膜電(diàn)位熒光(guāng)探針

産品關鍵詞：

DiOC2(3)；DiOC6(3)；壬基吖啶橙NAO；JC-10 線粒體膜電(diàn)位探針；MitoTracker® Green FM 綠色線粒體熒光(guāng)探針；羅丹明123 Rhodamine 123；CAS NO：905-96-4；

産品信息

【溫馨提示】：見(jiàn)我司懋康生(shēng)物(wù)（maokangbio）提供的線粒體熒光(guāng)探針産品專題。

産品描述

DiOC2(3)，英文全名3,3'-Diethyloxacarbocyanine Iodide，CAS NO. 905-96-4，是一(yī)種膜電(diàn)位探針，可通(tōng)過流式細胞儀檢測根據發射波長(cháng)熒光(guāng)信号比來分析細菌活力。DiOC2(3)常用來測定細菌膜電(diàn)位，也能(néng)用于研究某些活細胞比如大鼠胚胎成纖維細胞、猴腎細胞、中國(guó)倉鼠肺成纖維細胞和小(xiǎo)鼠3T3成纖維細胞。

DiOC2(3)用來檢測細菌膜電(diàn)位的工(gōng)作原理在于：DiOC2(3)在所有細菌細胞内發綠色熒光(guāng)，由于更高(gāo)的膜電(diàn)位引起染料分子發生(shēng)自(zì)聚作用，使得染料的熒光(guāng)往紅(hóng)色發射波長(cháng)處遷移，紅(hóng)色熒光(guāng)強度增高(gāo)，此時紅(hóng)色/綠色熒光(guāng)信号比高(gāo)。質子離子載體比如CCCP，通(tōng)過去除質子梯度來破壞線粒體膜電(diàn)位，從(cóng)而導緻紅(hóng)色熒光(guāng)強度降低(dī)，此時紅(hóng)色/綠色熒光(guāng)信号比降低(dī)。DiOC2(3)的最大激發和發射波長(cháng)約482nm和497nm。經DiOC2(3)标記的細胞用流式細胞儀分析（用488nm激發，分别用适合FITC（綠色）和Texas Red（紅(hóng)色）的發射濾片來檢測）。

産品特性

1）   CAS NO.：905-96-4

2）   化學名：3-ethyl-2-[3-(3-ethyl-2(3H)-benzoxazolylidene)-1-propenyl]-benzoxazolium iodide

3）   同義名：3,3'-Diethyloxacarbocyanine Iodide 3,3′-二乙基氧雜(zá)羰花青碘; DiOC2(3) Iodide; DOC Iodide;

4）   分子式：C21H21IN2O2

5）   分子量：460.31

6）   純度：>95.0% (HPLC)

7）   Ex/Em：482/497nm（in methanol）

8）   外觀：紅(hóng)色粉末或結晶

9）   溶解性：溶于DMSO，DMF，無水(shuǐ)乙醇，微溶于水(shuǐ)

10）化學結構式：

保存與運輸方法

保存：-20℃避光(guāng)幹燥保存，至少一(yī)年(nián)有效。

運輸：室溫運輸。

注意事(shì)項

1）   熒光(guāng)染料均存在淬滅問題，請盡量注意避光(guāng)，以減緩熒光(guāng)淬滅。

2）   為(wèi)了您的安全和健康，請穿實驗服并戴一(yī)次性手套操作。

使用方法

1.染色液制備

1）儲存液制備：用 DMSO或無水(shuǐ)乙醇配置濃度1～5 mM的儲存液。例如，取10mg DiOC2(3)（Mw：460.31 g/mol）溶于4.35ml 無水(shuǐ)DMSO中，充分溶解，即得到(dào)5mM的儲存液。【注意】未使用的儲存液分裝儲存在-20℃，避免反複凍融，且要避光(guāng)保存。

2）工(gōng)作液制備：用合适的緩沖液（如：無血清培養基，HBSS或PBS）稀釋儲存液，調整到(dào)合适的工(gōng)作濃度。【注意】：工(gōng)作液最終濃度需要根據不同細胞系和實驗體系來優化。

2.染色方法

對于細菌染色，最好取對數生(shēng)長(cháng)期的健康細菌，用無菌PBS或其他适當平衡鹽緩沖液稀釋到(dào)~1 x 106cells/ml。也可直接從(cóng)細菌培養液中取适量的細菌不經清洗進行稀釋。細菌染色用DiOC2(3)的工(gōng)作濃度可為(wèi)30μM，室溫染色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.

相(xiàng)關産品

參考文獻

[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【版權歸MKBio懋康所有】

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