20S蛋白酶體測定試劑盒| 20S Proteasome Assay Kit 貨號Abnova™ KA1352

蛋白酶體(proteasome)是一種多催化蛋白酶複合物(multicatalytic proteinase complex )，參與選擇性降解細胞內蛋白質。 20S蛋白酶體( 20S proteasome)是大蛋白降解的蛋白水解核心顆粒複雜的26S蛋白酶體( 26S proteasome)。這種 700 kDa 的蛋白質普遍分佈在細胞質和細胞質中。所有真核生物的細胞核。 該蛋白質負責降解許多細胞週期控制因子(cell-cycle control factors)、信號轉導因子(signal transduction factors)、轉錄因子( transcription factors)以及癌基因(oncogene)和抗癌基因產物(anti-oncogene products)，從而控制細胞增殖(cell proliferation)、分化( differentiation)和凋亡(apoptosis)。 20S蛋白酶體( 20S proteasome)表達的變化記錄在許多生理和病理條件下，例如骨骼肌萎縮(skeletal muscle atrophy )和阿爾茨海默神經變性(Alzheimer neurodegeneration)。乙醇抑制蛋白酶體活性(Ethanol inhibits proteasome activity)，導致不溶性物質積累被稱為馬洛里體的蛋白質聚集體，是酒精性肝病(alcoholic liver diseases)的常見特徵。 另一方面，蛋白酶體抑制劑表現出抗炎(anti-inflammatory )和抗增殖作用(antiproliferative effects)，證據表明蛋白酶體可能是治療癌症和炎症性疾病的重要藥物靶點。

測定原理 (Principle of the Assay)

20S 蛋白酶體檢測試劑盒(The 20S Proteasome Assay Kit) 採用特定的 20S 底物 SUC-LLVY-AMC，在裂解後活性酶產生高螢光產物，可以使用激發和發射進行測量波長分別為360 nm和480 nm。該套件易於使用，可輕鬆適應高調節 20S 蛋白酶體激活的治療化合物的通量篩選。jurkat cell lysate supernatant包含含有高水平 20S 活性的裂解物上清液，用作陽性對照。試劑盒中還包含用於證明特異性的的基材一種特定的 20S 抑製劑表沒食子兒茶素沒食子酸酯 (epigallocatechin gallate, EGCG)。

20S Proteasome Activity in Jurkat Cell Lysate Supernatant.

Figure 1: 20S Proteasome Activity in Jurkat Cell Lysate Supernatant. Jurkat cells were seeded in a 96-well plate in 100 μL of culture medium at a density indicated on the graph.
Cells were then processed for measurement of 20S activity according to the protocol described in the Assay Procedure section. Relative fluorescent intensity was then measured with a plate reader.
Note: Lactacystin is a known selective 20S inhibitor.

 

Figure 3: H2O2 inhibits 20S Proteasome Activity in Raw 264.7 cells

Raw 264.7 cells were seeded in a 96-well plate in 100 μL of culture medium at 8 x 104 cells/well and grown in DMEM containing 10% FBS for three days. Cells were then treated with different doses of H2O2 as indicated on the chart for 30 minutes and processed for measurement of 20S activity according to the protocol described in the Performing the Assay section. Relative fluorescent intensity was then measured with a plate reader.

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