單線态氧熒光(guāng)探針 Singlet Oxygen Sensor Green（SOSG）

産品關鍵詞：

Singlet Oxygen Sensor Green®, SOSG；單線态氧（Singlet Oxygen,1O2）；單線态氧熒光(guāng)探針；S36002；Thermo Fisher; ROS（Reactive oxygen species）活性氧物(wù)質；

活性氧引起的許多(duō)生(shēng)理損傷都可能(néng)歸因于單線态氧（1O2），包括與脫氧鳥苷選擇性反應形成8-羟基脫氧鳥苷（8-OHdG）的核酸修飾。單線态氧的壽命足夠長(cháng)（水(shuǐ)中有4.4μs）使其能(néng)夠充分擴散進入細胞和組織。實驗室中通(tōng)常用三種之一(yī)的方法來生(shēng)成單線态氧：1）使用感光(guāng)染料對二氧（3O2）進行光(guāng)化學反應來制備；2）通(tōng)過對一(yī)種過氧化物(wù)或二氧環丁烷進行加熱分解制備；3）氧氣流内通(tōng)過微波放(fàng)電(diàn)來制備；單線态氧通(tōng)過在1270nm處具弱化學發光(guāng)的典型特征直接被檢測。

Thermo fisher（molecular probe）公司開(kāi)發的Singlet Oxygen Sensor Green®, SOSG是目前商業(yè)化可得且應用最廣泛的單線态氧熒光(guāng)探針，從(cóng)材料學到(dào)藥物(wù)學領域，比如光(guāng)動力學療法。

【特異性強】

Singlet Oxygen Sensor Green®, SOSG高(gāo)度選擇性結合單線态氧（1O2）。不像其他的熒光(guāng)和化學發光(guāng)的單線态氧檢測試劑，SOSG與其他活性氧物(wù)質（ROS），包括羟基自(zì)由基（·OH）、超氧陰離子（·O2–）和一(yī)氧化氮（NO）無可見(jiàn)反應【見(jiàn)圖1】。

圖1、Singlet Oxygen Sensor Green (SOSG)的熒光(guāng)反應和對單線态氧（1O2）的特異性。A）用熒光(guāng)光(guāng)度計（Ex/Em= 488/525nm）測定溶液内的熒光(guāng)，溶液内含SOSG（1 µM ）和亞甲基藍（10 µM）溶于100mM pH 7.5 Tris Buffer; 單線态氧清除劑疊氮鈉（NaN3，1 mM）；或50% D2O（重水(shuǐ)，能(néng)夠延長(cháng)1O2的壽命）。每20s完成一(yī)次熒光(guāng)測定，每次測定間隔30s（如灰條标示）。在這30s的間隔中，樣本暴露于激光(guāng)輻射（630–680 nm，<5 mW），導緻亞甲基藍光(guāng)敏生(shēng)成1O2。B）用熒光(guāng)光(guāng)度計（Ex/Em=488/525nm）測定溶液内的熒光(guāng)，溶液為(wèi)50mM pH 7 Tris buffer，1mM 黃嘌呤，SOSG（1 µM ），或含SOSG（1 µM ），或含二氫羅丹明123。約20s之後，加入50 mU/mL黃嘌呤氧化酶（XO）。XO催化黃嘌呤的氧化，産生(shēng)尿酸和超氧化物(wù)。超氧化物(wù)瞬間被H2O2降解。

【熒光(guāng)特征】

Singlet Oxygen Sensor Green®, SOSG與單線态氧（1O2）反應前，探針最初發弱藍色熒光(guāng)，激發峰在372nm和393nm，發射峰在395nm和416nm。當含單線态氧（1O2），探針發射綠色熒光(guāng)，光(guāng)譜特征類似于熒光(guāng)素（最大激發/發射波長(cháng)約504/525nm）。

【線性化關系】

研究發現Singlet Oxygen Sensor Green®, SOSG的熒光(guāng)産物(wù)在某些溶液中随著(zhe)時間被降解，且在不含單線态氧的堿性pH溶液中，Singlet Oxygen Sensor Green®, SOSG會(huì)發熒光(guāng)。不過，當進行适當的操作，綠色熒光(guāng)信号的強度與單線态氧（1O2）含量相(xiàng)關，且不會(huì)受其他ROS的明顯幹擾。

【應用】

Singlet Oxygen Sensor Green®, SOSG證實能(néng)用來檢測溶液和植物(wù)組織中的單線态氧（1O2）。

【使用方法】

1.       SOSG儲存液制備：收到(dào)貨按照(zhào)≤-20℃避光(guāng)幹燥保存。用甲醇來制備儲存液，每100µg粉末加入33µl甲醇制備~5mM儲存液。正式實驗前制備工(gōng)作液，不能(néng)用完的工(gōng)作液需丢棄，不可保存後再用。

2.       溶液檢測：SOSG不具細胞膜滲透性，設計用來檢測溶液環境内的單線态氧。最佳的稀釋緩沖液和工(gōng)作濃度需根據經驗來調整。建議起始工(gōng)作濃度為(wèi)1-10µM。

【使用方法-來自(zì)文獻】

一(yī)、文獻來源：Flors C et al. Imaging the production of singlet oxygen in vivo using a new fluorescent sensor, Singlet Oxygen Sensor Green. J Exp Bot. 2006;57(8):1725-34. Epub 2006 Apr 4.

圖2. 肌紅(hóng)蛋白(bái)加入脂質體後随著(zhe)時間的SOSG熒光(guāng)變化（Changes in SOSG fluorescence with time after the addition of myoglobin to liposomes）。Singlet oxygen sensor green (SOSG) reagent was dissolved in methanol to make a stock solution of 500 μM. SOSG (final concentration 50 μM) was added to liposomes (2 mg ml−1) in 50 mM sodium phosphate buffer pH 7.4 in a Petri dish (2 cm diameter). Lipid oxidation was started by the addition of ferric horse myoglobin (10 μM). Following excitation of SOSG at 480 nm using a 75 W xenon lamp (Leica UK Ltd, Milton Keynes, UK) with a 480 nm interference filter and heat filter, images of the fluorescence from SOSG in the samples were produced using a Peltier-cooled (200 K) charge-coupled device (CCD) camera (576×384 pixels, Wright Instruments, Enfield, UK) with a 532 nm interference filter in the objective.

圖3. 拟南(nán)芥葉子内SOSG熒光(guāng)産生(shēng)的動力學變化（Kinetics of the development of SOSG fluorescence in Arabidopsis leaves）。Leaves were grown at a PPFD of 200 μmol m−2 s−1, detached and infiltrated with SOSG, and then the top halves of the leaves exposed to a PPFD of 650 μmol m−2 s−1 (high light). The bottom halves were kept in the dark; images were taken after 0, 5, 10, 20, 30, and 50 min.

二、文獻來源：Chen T et al. Singlet Oxygen Plays an Essential Role in the Root’s Response to Osmotic Stress. Plant Physiol. 2018 Aug;177(4):1717-1727.

圖4. PEG處理後單線态氧和碘化丙啶細胞滲透性的變化圖（Appearance of singlet oxygen and changes in cell permeability to propidium iodide after PEG treatment）。Root tips of 7-d-old seedlings stained with propidium iodide (PI) and singlet oxygen sensor green (SOSG). Median confocal slices are shown for each of these stains, along with the corresponding bright field (BF) image. Treatments were for the times indicated on the right: no treatment; 1 to 2 h, early; 3 to 4 h, mid; 5 to 8 h, late. Zones are as follows: AM, apical meristem; BM, basal meristem; and EZ, elongation zone. The median plane of the root was imaged where opaque regions in PI staining are indicative of cell death. White outlined arrows indicate SOSG and PI staining of live cells (i.e. the stain is observed in the perimeter of the cell). Solid white arrows indicate SOSG and PI staining of dead cells (i.e. where the PI stain permeates and fills the cell). Graphs on the left show quantification of the fluorescence, calculated by estimating the area of the root (using threshold above background) in a median slice that displayed SOSG or opaque PI fluorescence (using threshold above that obtained from live cells). Scale bar, 100 µm. For quantification, error bars are se, n ≥ 8.

【争議點】

Singlet Oxygen Sensor Green®, SOSG不具有細胞膜滲透性，能(néng)用于細胞單線态氧（1O2）檢測？

專利持有方thermo（molecular probe）官方說明此探針不具有細胞膜滲透性，适用于溶液體系單線态氧（1O2）檢測。然而，Gollmer A et al.報(bào)道SOSG能(néng)插入活哺乳動物(wù)細胞（文獻來源：Gollmer A et al. Singlet Oxygen Sensor Green®: photochemical behavior in solution and in a mammalian cell. Photochem Photobiol. 2011 May-Jun;87(3):671-9.）。

訂購信息：Thermo fisher原裝正品，大包裝拆開(kāi)單賣，100μg /支，單支特價700元。懋康生(shēng)物(wù)長(cháng)期現貨供應，歡迎咨詢，QQ:2971634497，或電(diàn)話：021-54736159。

【溫馨提示】：見(jiàn)我司懋康生(shēng)物(wù)整理的活性氧檢測探針專題（Probes for reactive oxygen species）。

相(xiàng)關産品：ROS研究常用的配套産品，需要了解更多(duō)信息歡迎來電(diàn)咨詢，021-54736159，QQ:2971634497。

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