Live or Dead 細(xì)胞活性檢測試劑盒是美國AAT Bioquest生產(chǎn)的細(xì)胞活性檢測試劑盒，AAT Bioquest的Mycolight 細(xì)菌活力測定試劑盒可在革蘭氏陽性和陰性細(xì)菌中提供細(xì)菌活力的雙色熒光測定。該試劑盒利用我們的綠色熒光核酸染色劑MycoLight Green和紅色熒光核酸染色碘化丙啶的混合物。單獨(dú)使用時，MycoLight 綠色污漬通常會標(biāo)記群體中的所有細(xì)菌（活的和死的）。相比之下，碘化丙錠只穿透膜受損的細(xì)菌，當(dāng)兩種染料都存在時，導(dǎo)致MycoLight 綠色染料熒光的減少。因此，使用MycoLight 綠色和碘化丙啶的適當(dāng)混合物，具有完整細(xì)胞膜的活細(xì)菌發(fā)出綠色熒光，而具有損傷膜的死亡或瀕死的細(xì)菌 產(chǎn)生紅色熒光。Mycolight 細(xì)菌活力測定試劑盒是監(jiān)測細(xì)菌種群活力的強(qiáng)大工具，可作為細(xì)胞膜完整性的函數(shù)。可以在510-530nm（FITC濾光片）和600-660nm（德克薩斯紅色濾光片光片）處以488nm處（常見的激發(fā)光源）激發(fā)熒光測定染色的細(xì)胞。百螢生物是AAT Bioquest的中國代理商，為您提供優(yōu)質(zhì)的Live or Dead 細(xì)胞活性檢測試劑盒。 

樣品實(shí)驗方案

簡要概述

1.用測試化合物制備細(xì)胞
2.添加染料加工溶液
3.在室溫或37°C孵育30分鐘至1小時
4.在Ex / Em = 610/650 nm（截止= 630 nm，紅色）和Ex / Em = 360/450 nm（截止= 420 nm，藍(lán)色）或在熒光顯微鏡下觀察（Texas red/Cy5通道   活細(xì)胞），（DAPI通道  死細(xì)胞）

工作溶液配制

將5μL的200X Cellbrite Red（組分A）和5μL的200X Nuclear Blue DCS1（組分C）加入1mL的測定緩沖液（組分B）中并充分混合以制備染料 - 工作溶液。 該染料加工溶液在室溫下穩(wěn)定至少1小時。 注意：由于染色條件可能因細(xì)胞類型的不同而異，因此建議單獨(dú)確定組分A和C的適當(dāng)濃度。有關(guān)細(xì)胞樣品制備的指南，請點(diǎn)擊查看。

操作步驟

1.根據(jù)標(biāo)準(zhǔn)方案準(zhǔn)備細(xì)胞。 注意：我們用星形孢菌素（SS）在37℃下處理HeLa細(xì)胞4小時以誘導(dǎo)細(xì)胞凋亡。 詳細(xì)信息請參見圖1。

2.用100μL/孔（96孔板）或25μL/孔（384孔板）的染料加工溶液替換生長培養(yǎng)基。

3.將染料加工溶液板在室溫或37°C孵育30分鐘至1小時，避光。

4.用HHBS，PBS或您選擇的緩沖液洗滌細(xì)胞兩次。

5.向細(xì)胞中加入100μL/孔（96孔板）或25μL/孔（384孔板）的測定緩沖液（組分B）。

6.用熒光顯微鏡監(jiān)測熒光信號，用德克薩斯紅或Cy5過濾器檢測活細(xì)胞，用DAPI過濾死細(xì)胞。 還可以使用熒光酶標(biāo)儀（底部讀取模式）在Ex / Em = 610 / 650nm（截止= 630nm，紅色）和Ex / Em = 360 / 450nm（截止= 420nm，藍(lán)色）下分析熒光強(qiáng)度。

數(shù)據(jù)分析

參考文獻(xiàn)

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