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當(dāng)前位置:深圳欣博盛生物科技有限公司>>技術(shù)文章>>組蛋白去乙酰化酶熒光活性檢測試劑盒介紹
ENZO熱銷產(chǎn)品——FLUOR DE LYS® HDAC fluorometric activity assay kit組蛋白去乙?;笩晒饣钚詸z測試劑盒
目前,,組蛋白去乙酰化酶抑制劑已經(jīng)顯示出其作為抗腫瘤藥物的前景,,這也自然激發(fā)了研究者們對(duì)于篩選HDAC(Histone deacetylase,,組蛋白去乙酰化酶)抑制化合物的興趣,。Enzo的FLUOR DE LYS®HDAC熒光活性檢測試劑盒(貨號(hào):BML-AK500-0001)是一種靈敏,、便捷的組蛋白去乙酰化酶檢測試劑盒,,可替代使用傳統(tǒng)的放射性標(biāo)記的乙?;M蛋白和肽/HPLC法檢測組蛋白去乙酰化酶的方案,。
FLUOR DE LYS®去乙?;笝z測平臺(tái)是Enzo的升級(jí)酶活性檢測系列產(chǎn)品的基礎(chǔ),其中的產(chǎn)品就包括用于篩選HDAC(組蛋白去乙?;福┗钚哉{(diào)節(jié)劑和Sirtuin(長壽蛋白,,一種去乙酰化酶)活性調(diào)節(jié)劑的試劑盒,。Enzo還可提供基于熒光共振能量轉(zhuǎn)移(FRET)的試劑盒,,用以檢測基質(zhì)金屬蛋白酶(MMPs)和天冬氨酸特異性半胱氨酸蛋白酶(Caspase)的活性,以及提供基于磷酸特異性抗體的激酶檢測試劑盒,。
這些檢測證明了Enzo在組裝小肽,、合成活性重組蛋白、熒光標(biāo)記和檢測技術(shù),,以及抗體開發(fā)等多方面協(xié)同的能力,,也保障了Enzo能夠生產(chǎn)出功能強(qiáng)大且重復(fù)性好的檢測試劑盒,。
Enzo的酶活性檢測試劑盒均具有以下特點(diǎn):
高品質(zhì):高活性、高純度的重組酶,,可呈現(xiàn)高質(zhì)量的底物結(jié)合效果
靈活性:批量酶和底物可用于篩選后研究
強(qiáng)創(chuàng)新:多種readout 格式的專-利底物/顯影劑化學(xué)物質(zhì)
作用機(jī)制:
FLUOR DE LYS® HDAC fluorometric activity assay kit基于Enzo特-有的 FLUOR DE LYS®(熒光法組蛋白去乙?;?賴氨酸)底物和顯影劑組合,只需2步簡單的混合步驟,,即可在同一個(gè)96 孔板上進(jìn)行檢測,。首先,將含有乙?;嚢彼醾?cè)鏈的FLUOR DE LYS®底物與含有HDAC活性的樣品(HeLa核或其他提取物,、純化酶、珠結(jié)合免疫復(fù)合物等)一起孵育,。底物的去乙?;瘯?huì)使底物增敏,,所以在第二步中,,將其與FLUOR DE LYS®顯影劑混合后會(huì)產(chǎn)生熒光團(tuán)。
該檢測方法目前已經(jīng)成功用于所有已知的I類HDAC——HDAC1,、HDAC2,、HDAC3和HDAC8(見產(chǎn)品數(shù)據(jù)表)和II 類HDAC 4-7、9和10,,以及人類Sir2同源物和SIRT1(見產(chǎn)品數(shù)據(jù)表)的制備,。Enzo Life Sciences的研究表明,F(xiàn)LUOR DE LYS®底物具有細(xì)胞滲透性,,并可在原位被細(xì)胞HDAC去乙?;C撘阴,;牡孜镌诩?xì)胞內(nèi)積聚后,,可通過向細(xì)胞裂解液中加入FLUOR DE LYS®顯影劑進(jìn)行定量測定。
Figure 1. HDAC熒光活性檢測的反應(yīng)方案。底物去乙?;蛊鋵?duì)顯影劑敏感,,從而產(chǎn)生熒光團(tuán)(symbol)。熒光團(tuán)可被360 nm的光激發(fā),,并發(fā)射出460 nm的光,,發(fā)射光在熒光平板閱讀器上可被檢測到。
產(chǎn)品特點(diǎn):
使用提供的核提取物,,有助于檢測裂解物,、免疫沉淀物以及篩選抑制劑,;
含有HeLa核提取物,作為HDAC 1和2的豐富來源,,可用作陽性對(duì)照,,也可用作篩選HDAC活性的來源;
與I & IIb類HDAC和Sirtuin兼容(添加NAD+),;
無放射性,,沒有提取步驟;
所提供的試劑足夠分析100-200次,。
產(chǎn)品信息:
產(chǎn)品貨號(hào) | BML-AK500-0001 |
產(chǎn)品名稱 | FLUOR DE LYS® HDAC fluorometric activity assay kit |
規(guī)格 | 1*96wells |
使用/穩(wěn)定性 | Store all components, except the microtiter plate, at -80℃ for the highest stability. The HeLa Nuclear Extract, BML-KI140, must be handled with particular care in order to retain maximum enzymatic activity. Defrost it quickly in a RT water bath or by rubbing between fingers, then immediately store on an ice bath. The remaining unused extract should be refrozen quickly, by placing at -80°C. If possible, snap freeze in liquid nitrogen or a dry ice/ethanol bath. To minimize the number of freeze/thaw cycles, aliquot the extract into separate tubes and store at -80℃. The FLUOR DE LYS® Substrate, BML-KI104, when diluted in Assay Buffer, may precipitate after freezing and thawing. It is best, therefore, to dilute only the amount needed to perform the assays of that day. |
運(yùn)輸 | Dry Ice |
長期保存 | -80℃ |
試劑盒組分 | Nuclear Extract from HeLa Cells (human cervical cancer cell line) (Prod. No. BML-KI140-0100) (100 µl; In 0.1M KCl, 20mM HEPES/NaOH, pH 7.9, 20% (v/v) glycerol, 0.2mM EDTA, 0.5mM DTT, 0.5Mm PMSF, prepared according to a modification of J.D. Dignam et al. (1983) and S.M. Abmayr et al. (1988)). Storage: -80℃, avoid freeze/thaw cycles FLUOR DE LYS® Substrate (Prod. No. BML-KI104-0050) (50 µl; 50mM in DMSO) Storage: -80℃ FLUOR DE LYS® Developer Concentrate (20×) (Prod. No. BML-KI105-0300) (300 µl; 20×stock solution, dilute in assay buffer before use) Storage: -80℃ Trichostatin A (HDAC Inhibitor) (Prod. No. BML-GR309-9090) (100 µl; 0.2mM in DMSO) Storage: -80℃ FLUOR DE LYS® Deacetylated Standard (Prod. No. BML-KI142-0030) (30 µl; 10mM in DMSO) Storage: -80℃ HDAC Assay Buffer (Prod. No. BML-KI143-0020) (20 ml; 50mM TRIS/Cl, pH 8.0, 137mM NaCl, 2.7mM KCl, 1mM MgCl2) Storage: -20℃ 1/2 volume microplate (Prod. No. BML-KI101) Storage: Ambient 1/2 volume white microplate (Prod. No. BML-K571) Storage: Ambient |
產(chǎn)品引用文獻(xiàn):
The activation of histone deacetylases 4 prevented endothelial dysfunction: A crucial mechanism of HuangqiGuizhiWuwu Decoction in improving microcirculation dysfunction in diabetes: M. Chen, et al.; J. Ethnopharmacol. 307, 116240 (2023), Abstract;
Four New Anthraquinones with Histone Deacetylase Inhibitory Activity from Ventilago denticulata Roots: N. Hangsamai, et al.; Molecules 27, 1088 (2022), Abstract;
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