| 励起波長 |
530
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| 蛍光波長 |
590
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| 検出系 |
蛍光
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| 溶媒 |
DMSO
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| 使用目的 |
カ ルシウムを通じて伝達する GPCRを発現する細胞は、細胞膜にクロスできるABD社独自の Rhod-4 NWによってpre-load されます。Rhod-4 は、HTS スクリーニングに利用できる赤色のカルシウムインジケーターです。いったん細胞内に入ると Quest Rhod-4TM の親油性ブロッキンググループが非特異的細胞エステラーゼによって開裂され、その結果マイナスにチャージした蛍光色素が細胞内にとどまり、カルシウムに結 合することで蛍光強度が増します。細胞がスクリーニングする化合物によって刺激を受けた際、レセプターが細胞内カルシウムの放出を伝達し、その結果、 Rhod-4 の蛍光がさらに増強します (Ex: 488, 514, 532, 546nm、Em: 〜555nm)。Quest Rhod-4TMは、カルシウムと複合体を形成すると、250倍以上の蛍光増強を示し、細胞内カルシウム測定のための理想的なインジケーターです。本キット はGタンパク質共役受容体(GPCRs) やカルシウムチャンネルを観察するための最適化したアッセイです。ほんアッセイは便利な96-well もしくは 384-well マクロタイタープレートフォーマットで、オートメーションにも対応しています。
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| 構成内容 |
・Rhod-4 NW 1 vial, lyophilized
・10X PluronicR F127 Plus 10 bottles (1 mL/bottle)
・HHBS 1 bottle (10 mL)
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| その他 |
[測定波長(nm)] Ex 530/Em 555
[内容･その他]Calcium flux assays are preferred methods in drug discovery for screening G protein coupled receptors (GPCR). Screen Quest･Rhod-4 NW Calcium Assay Kitprovides a homogeneous fluorescence-based assay for detecting the intracellular calcium mobilization. Cells expressing a GPCR of interest that signals through calcium are pre-loaded with our proprietary Rhod-4 NW which can cross cell membrane. Rhod-4is the brightest red calcium indicator available for HTS screening. Once inside the cell, the lipophilic blocking groups of Quest Rhod-4･are cleaved by non-specific cell esterase, resulting in a negatively charged fluorescent dye that stays inside cells, and its fluorescence is greatly enhanced upon binding to calcium. When cells stimulated with screening compounds, the receptor signals release of intracellular calcium, which greatly increase the fluorescence of Rhod-4. The characteristics of its long wavelength, high sensitivity, and >250 times fluorescence increases (when it forms complexes with calcium) make Quest Rhod-4･an ideal indicator for measurement of cellular calcium. This Screen Quest Rhod-4 NW Calcium Assay Kit provides an optimized assay method for monitoring G-protein-coupled receptors (GPCRs) and calcium channels. The assay can be performed in a convenient 96-well or 384-well microtiter-plate format and easily adapted to automation.
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| 参考文献 |
Territo PR, Heil J, Bose S, Evans FJ, Balaban RS.. (2007). Fluorescence absorbance inner-filter decomposition: the role of emission shape on estimates of free Ca(2+) using Rhod-2. Appl Spectrosc, 61, 138.
Stamm C, del Nido PJ.. (2004). Protein kinase C and myocardial calcium handling during ischemia and reperfusion: lessons learned using Rhod-2 spectrofluorometry. Thorac Cardiovasc Surg, 52, 127.
Bednar B, Cunningham ME, Kiss L, Cheng G, McCauley JA, Liverton NJ, Koblan KS.. (2004). Kinetic characterization of novel NR2B antagonists using fluorescence detection of calcium flux. J Neurosci Methods, 137, 247.
Martin VV, Beierlein M, Morgan JL, Rothe A, Gee KR.. (2004). Novel fluo-4 analogs for fluorescent calcium measurements. Cell Calcium, 36, 509.
Stamm C, Friehs I, Choi YH, Zurakowski D, McGowan FX, del Nido PJ.. (2003). Cytosolic calcium in the ischemic rabbit heart: assessment by pH- and temperature-adjusted rhod-2 spectrofluorometry. Cardiovasc Res, 59, 695.
Lannergren J, Westerblad H, Bruton JD.. (2001). Changes in mitochondrial Ca2+ detected with Rhod-2 in single frog and mouse skeletal muscle fibres during and after repeated tetanic contractions. J Muscle Res Cell Motil, 22, 265.
Du C, MacGowan GA, Farkas DL, Koretsky AP.. (2001). Calibration of the calcium dissociation constant of Rhod(2)in the perfused mouse heart using manganese quenching. Cell Calcium, 29, 217.
MacGowan GA, Du C, Glonty V, Suhan JP, Koretsky AP, Farkas DL.. (2001). Rhod-2 based measurements of intracellular calcium in the perfused mouse heart: cellular and subcellular localization and response to positive inotropy. J Biomed Opt, 6, 23.
Du C, MacGowan GA, Farkas DL, Koretsky AP.. (2001). Calcium measurements in perfused mouse heart: quantitating fluorescence and absorbance of Rhod-2 by application of photon migration theory. Biophys J, 80, 549.
Muriel MP, Lambeng N, Darios F, Michel PP, Hirsch EC, Agid Y, Ruberg M.. (2000). Mitochondrial free calcium levels (Rhod-2 fluorescence) and ultrastructural alterations in neuronally differentiated PC12 cells during ceramide-dependent cell death. J Comp Neurol, 426, 297.
do Ceu Monteiro M, Sansonetty F, Goncalves MJ, O'Connor JE.. (1999). Flow cytometric kinetic assay of calcium mobilization in whole blood platelets using Fluo-3 and CD41. Cytometry, 35, 302.
Cheng H, Song LS, Shirokova N, Gonzalez A, Lakatta EG, Rios E, Stern MD.. (1999). Amplitude distribution of calcium sparks in confocal images: theory and studies with an automatic detection method. Biophys J, 76, 606.
Smith GD, Keizer JE, Stern MD, Lederer WJ, Cheng H.. (1998). A simple numerical model of calcium spark formation and detection in cardiac myocytes. Biophys J, 75, 15.
Perez-Terzic C, Stehno-Bittel L, Clapham DE.. (1997). Nucleoplasmic and cytoplasmic differences in the fluorescence properties of the calcium indicator Fluo-3. Cell Calcium, 21, 275.
Tretyn A, Kado RT, Kendrick RE.. (1997). Loading and localization of Fluo-3 and Fluo-3/AM calcium indicators in sinapis alba root tissue. Folia Histochem Cytobiol, 35, 41.
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| 使用文献 |
Territo PR Heil J, Bose S, Evans FJ, Balaban RS. (2007). Fluorescence absorbance inner-filter decomposition: the role of emission shape on estimates of free Ca(2+) using Rhod-2. Appl Spectrosc, 138.
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| [注意事項] |
湿気を防ぎ､遮光して保存
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※サムネイル画像をクリックすると拡大画像が表示されます。
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| メーカー |
品番 |
包装 |
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ABD
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36330
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1 KIT
[1 Plate]
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※表示価格について
| 当社在庫 |
なし
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| 納期目安 |
1週間程度
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| 保存温度 |
-２０℃
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