脂肪組織/培養之脂肪細胞蛋白質萃取套組 |Invent貨號AT-022

產品名稱: Minute™  脂肪組織/培養之脂肪細胞蛋白質萃取套組| Adipose Tissues/Cultured Adipocyte for Adipose Tissues/Cultured Adipocytes| 貨號AT-022

產品介紹: 脂肪組織特別是白色脂肪組織（WAT）已經被認為是除了其能量存儲功能之外的重要的內分泌和炎症器官。來自脂肪組織的蛋白質的分離與分析對於理解許多生理/病理狀況越來越重要。然而，由於其高脂質和低蛋白質含量，讓WAT和棕色脂肪組織（BAT）的分離和分析在技術上是非常具有挑戰性的。存在於生物樣品中的水 – 油乳液難以分離；Invent開發新技術，具有獨特的表面性質和預定的孔徑和厚度的多孔過濾器與特別配製的不含洗滌劑的提取緩衝液結合，用於快速和有效地分離得自脂肪組織勻漿的水 – 油乳液。提取緩衝液具有比脂肪組織中的油更低的凝固點，並且水相可以通過使組織勻漿通過過濾器而從油相快速分離。與其他方法相比，提取高濃度蛋白質（2-3mg / ml）含量。可應用於2D gel analysis、 ELISA、 SDS-PAGE、, immunoblotting, immunoprecipitation、 enzyme activity assays、等分析。

Minute™蛋白萃取套組，使用創新『Column Filter』技術，效果更佳，完成只需 一 分鐘蛋白萃取只需要vortex、transfer、spin 。Minute ™一系列蛋白質提取試劑盒；適用於多種物種（包括哺乳動物、海洋動物、昆蟲、植物和細菌）快速和容易地提取/分離蛋白質的優異工具 。與市場上的任何其他蛋白質提取試劑盒不同，優化的緩衝液系統，結合最新的專有濾筒技術（spin column）；快速提高蛋白萃取的效能及方便性；用於總蛋白提取，細胞分離和膜蛋白分離，只需簡單四步驟即可完成蛋白質萃取。無須長時間等待、免除重複性的wash步驟。

SDS-PAGE (10%) profiles of total protein extracted from different adipose tissues. Lane 1, porcine WAT; Lane 2, chicken BAT; Lane 3, rat WAT. B. Western blottings of extracted proteins from rat WAT. Proteins were separated in 8-16% gradient SDS-PAGE and probed with following cellular protein marker antibodies:Anti-Na/K ATPase alpha1, a plasma membrane marker(Upstate, clone 464.6), anti-lamin B1, a nuclear envelope marker (ab16048, abcam Cambridge, MA), anti-ubiquinol-cytochrome C reductase core protein (abcam, ab 96333) and GAPDH, a cytosolic marker (Sigma). The specific protein bands were visualized by a substrate Opti-4CN (Bio-RAD).

注意事項: 

蛋白質最佳化，可使用protease inhibitors蛋白酶抑製劑。如果下游應用是需要比較久時間；或蛋白質提取物儲存長時間才使用，可於緩衝液A中加入protease inhibitors蛋白酶抑製劑使用。蛋白質濃度定量可應用BCA方法測定。 於蛋白質磷酸化研究，在使用前加入磷酸酶抑製劑cocktails of phosphatase inhibitors放緩衝液A中使用。**如果緩衝液B在較低溫度下存在沉澱物，37℃回溫直到沉澱完全溶解。

提取的蛋白質看起來略微模糊不清存在水不溶性細胞成分。它可以稀釋並直接用於ELISA檢測水溶性蛋白質。它也可以重懸於緩衝液B ( buffer B) 或緩衝液C ( buffer C)中，以溶解水不溶性蛋白質 (water insoluble proteins ) 並用於下游應用：
A.提取的蛋白質溶液中加入1/10的緩衝液B ( buffer B)，得到變性的蛋白質溶液（適用於SDS-PAGE，Westerns和其他應用）
B.提取的蛋白質溶液中加入1/10的緩衝液C ( buffer B)，得到未變性的蛋白質（適用於IP，ELISA和其他應用）
C.溶於2×2D凝膠樣品緩衝液中進行2D凝膠分析

注意：緩衝液A包含可能干擾質譜分析的組分。如果提取蛋白質用於MS分析，首先針對兼容緩衝液進行透析。另一種選擇是沉澱蛋白質。使用標準TCA蛋白沉澱方案。

引用文獻

Minute^TM Total Protein Extraction Kit for Adipose Tissue/Cultured cells (AT-022)

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