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Cell Meter 活細胞ATP檢測試劑盒

貨號23015存儲條件 在零下15度以下保存, 避免光照
規格100 Tests價格4140
Ex (nm)Em (nm)
分子量溶劑
產品詳細介紹


簡要概述

三磷酸腺苷(ATP)在細胞能量、代謝調節和細胞信號傳導中起著基本作用。它被稱為細胞內能量轉移的“貨幣分子單位”,以驅動活細胞中的許多生物過程和化學合成。 ATP也是細胞通訊的信號分子,在DNA和RNA合成中起著重要作用。它位于發生細胞呼吸的線粒體中。 ATP水平可用于測量細胞增殖和細胞周期動態。 Cell Meter 活細胞ATP檢測試劑盒使研究人員可以使用檢測細胞ATP的滲透型紅色熒光成像探針ATP Red 來檢測活細胞中的ATP水平。 ATP Red 旨在檢測活細胞線粒體中的ATP濃度。該探針與AMP,ADP,CMP,CDP,CTP,UMP,UDP,UTP,GMP,GDP或GTP的交叉反應性最小。

 

適用儀器


熒光酶標儀  
激發: Cy3/TRITC濾波片組
發射: Cy3/TRITC濾波片組
推薦孔板: 黑色透明孔板


產品說明書

實驗方案

概述

在生長培養基中準備細胞
將細胞與ATP Red 工作溶液在37°C下孵育15-30分鐘
去掉ATP Red 工作溶液
使用Cy3 / TRITC濾波片組的熒光顯微鏡進行檢測

 

準備細胞

對于貼壁細胞
在96孔板的生長培養基中以10,000至40,000個細胞/孔/ 90μL平板培養細胞過夜,而對于384孔板則以2,500至10,000個細胞/孔/ 20μL平板培養過夜。

對于懸浮細胞
離心培養液中的細胞,然后將細胞沉淀以50,000-100,000細胞/孔/ 90 µL的濃度懸浮于96孔聚D賴氨酸平板中,或10,000-25,000細胞/孔/ 20 / L以獲得384孔中的懸浮液 聚-D賴氨酸板。 實驗前,將板以800 rpm的速度離心2分鐘。
注意:應該對每個細胞系進行單獨評估,以確定最佳細胞密度。

 

溶液配制

除非另有說明,否則所有未使用的儲備溶液應分為一次性使用的等分試樣,并在制備后儲存在-20°C下。 避免重復凍融循環。

ATP Red 儲備溶液配制(500X)
將50 µL DMSO(組分C)添加到小瓶ATP Red (組分A)中,制成500X儲備液。
注意:50 µL的500X ATP Red 儲備溶液足以容納一個96孔板。 可以將未使用的ATP Red 儲備溶液分裝,并在≤-20°C下以較小的等分試樣存儲。 避光,并避免重復的凍融循環。

 

ATP Red 工作溶液配制
將5 µL 500X儲備溶液(組分A)添加到1 mL細胞培養基中,并充分混合。
注意:ATP Red 探針與我們檢測的大多數細胞系的細胞培養基兼容。 可以根據特定的細胞類型修改染色條件。

 

操作步驟

1.在生長培養基中準備細胞。
2.在細胞板中加入等體積的[100 µL /孔(96孔板)或25 µL /孔(384孔板)]。
注意:ATP Red 的最佳濃度因具體應用而異。
3.在避光的條件下,將細胞在37°C下孵育15-30分鐘。
4.從每個孔中除去工作溶液。 用測定緩沖液(組分B)或您選擇的緩沖液洗滌細胞兩次。
5.使用帶有Cy3 / TRITC濾光片組的熒光顯微鏡觀察細胞中的熒光信號。

 

圖示

圖1. HeLa細胞與ATP Red 和MitoLite Green FM共染色的熒光圖像在96孔黑色透明底板。 HeLa細胞用ATP Red 染色15分鐘,然后與100 nM MitoLite Green FM(Cat#22695)孵育30分鐘。 成像前用測定緩沖液洗滌兩次。 ATP Red 和MitoLite ATP Red 信號重疊良好。使用帶有Cy3 / TRITC和FITC濾光片的熒光顯微鏡對細胞成像。

 

參考文獻

Metabolic co-dependence of the oocyte and cumulus cells: essential role in determining oocyte developmental competence.
Authors: Richani, Dulama and Dunning, Kylie R and Thompson, Jeremy G and Gilchrist, Robert B
Journal: Human reproduction update (2021): 27-47

Mitochondrial C3a Receptor Activation in Oxidatively Stressed Epithelial Cells Reduces Mitochondrial Respiration and Metabolism.
Authors: Ishii, Masaaki and Beeson, Gyda and Beeson, Craig and Rohrer, Bärbel
Journal: Frontiers in immunology (2021): 628062

Cell-penetrating and mitochondrion-targeting molecules.
Authors: Appiah Kubi, George and Pei, Dehua
Journal: Methods in enzymology (2020): 311-328

Combinatorial roles of mitochondria and cGMP/PKG pathway in the generation of neuronal free Zn2+ under the presence of nitric oxide.
Authors: Yang, De-Ming and Huang, Chien-Chang and Chang, Yu-Fen
Journal: Journal of the Chinese Medical Association : JCMA (2020): 357-366

Dynamic regulation of subcellular mitochondrial position for localized metabolite levels.
Authors: Alshaabi, Haya and Heininger, Meara and Cunniff, Brian
Journal: Journal of biochemistry (2020): 109-117

Engineering a Reversible Fluorescent Probe for Real-Time Live-Cell Imaging and Quantification of Mitochondrial ATP.
Authors: Ren, Tian-Bing and Wen, Si-Yu and Wang, Lu and Lu, Peng and Xiong, Bin and Yuan, Lin and Zhang, Xiao-Bing
Journal: Analytical chemistry (2020): 4681-4688

Heat stress induces apoptosis through disruption of dynamic mitochondrial networks in dairy cow mammary epithelial cells.
Authors: Chen, Kun-Lin and Wang, Hui-Li and Jiang, Lin-Zheng and Qian, Yong and Yang, Cai-Xia and Chang, Wei-Wei and Zhong, Ji-Feng and Xing, Guang-Dong
Journal: In vitro cellular & developmental biology. Animal (2020): 322-331

Intracellular ATP levels influence cell fates in Dictyostelium discoideum differentiation.
Authors: Hiraoka, Haruka and Nakano, Tadashi and Kuwana, Satoshi and Fukuzawa, Masashi and Hirano, Yasuhiro and Ueda, Masahiro and Haraguchi, Tokuko and Hiraoka, Yasushi
Journal: Genes to cells : devoted to molecular & cellular mechanisms (2020): 312-326

Measurement of ATP concentrations in mitochondria of living cells using luminescence and fluorescence approaches.
Authors: Morciano, Giampaolo and Imamura, Hiromi and Patergnani, Simone and Pedriali, Gaia and Giorgi, Carlotta and Pinton, Paolo
Journal: Methods in cell biology (2020): 199-219

Mitochondria transfer enhances proliferation, migration, and osteogenic differentiation of bone marrow mesenchymal stem cell and promotes bone defect healing.
Authors: Guo, Yusi and Chi, Xiaopei and Wang, Yifan and Heng, Boon Chin and Wei, Yan and Zhang, Xuehui and Zhao, Han and Yin, Ying and Deng, Xuliang
Journal: Stem cell research & therapy (2020): 245



說明書
Cell Meter 活細胞ATP檢測試劑盒.pdf


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