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MKbio Forskolin 佛司可林/毛喉素(最經典的腺苷酸環化酶活化劑)
時間:2018-01-25     作者:懋康原創     文章來源:懋康生物    

Forskolin 佛司可林/毛喉素


產品標簽

Forskolin (Fsk) 佛司可林;Adenylate cyclase (AC) 腺苷酸環化酶;cAMP 環磷酸腺苷;Neuron Differentiation 神經元分化;CAS NO:66575-29-9;


景描述

環磷酸腺苷(cAMP)由腺苷酸環化酶(Adenylate cyclase , AC)(一種12-跨膜域酶)產生,AC催化ATP轉化生成3′,5′-cAMP和焦磷酸鹽。cAMP是一種重要的信號載體,在細胞對激素,神經遞體和其他細胞外信號的恰當的生理反應中是不可或缺的。環磷酸腺苷(cAMP)由AC合成且經磷酸二酯酶(phosphodiesterases, PDEs)降解。局部cAMP信號通過靶向信號組分到亞細胞區室和組裝信號復合物來獲得。 


產品描述

佛司可林(Forskolin,CAS NO.:66575-29-9)是一種來源于印度彩葉植物毛喉鞘蕊花(Coleus forskohlii)的天然生成二萜。通過作用于催化亞基直接激活腺苷酸環化酶(AC),引起細胞內cAMP水平的提高[1-2]。在穩定轉染且表達I型腺苷酸環化酶(AC1)的Sf9細胞,Forskolin與膜上的AC1結合(IC50=41nM)并活化其功能(EC50=0.5 μM,評估從ATP生成cAMP的激活實驗)[3]。Forskolin以濃度依賴性的方式發揮平滑肌松弛劑和血管擴張劑的功能,同時影響細胞質鈣離子水平和收縮成分的鈣離子敏感性[4]。Forskolin能夠與葡萄糖轉運體和某些離子通道相互作用,可用來檢測腺苷酸環化酶的表達、調控和G蛋白信號通路[5]。Forskolin用于幾種分化方案中以增強神經元分化能力[6-7]。


我司提供兩種形式的佛司可林(Forskolin):

一種是溶于DMSO的50mM儲存液形式,對應貨號是:MZ2301-50UL(~1mg),MZ2301-250UL(~5mg),直接用培養基稀釋到所需工作濃度即可。操作簡單方便。

一種是凍干粉形式,對應貨號是:MZ2301-50MG,MZ2301-100MG。需要先用DMSO或無水乙醇配制成儲存液,再用培養基稀釋到所需工作濃度使用。


Forskolin通常用在各種類型的完整細胞或組織培養物中來提高cAMP水平,常用工作濃度為10-50μM。具體的最佳工作濃度請參考文獻,或根據自身實驗體系,培養細胞和組織類型,通過實驗進行摸索和優化。



產品應用(T淋巴細胞使用Forskolin抑制IL-2合成的應用示例)

細胞類型:Jurkat cells (淋巴細胞樣T細胞系)

作用機制:Jurkat細胞能生成IL-2。正常生長情況,Jurkat細胞幾乎沒有或不生成IL-2。PMA,通過刺激PKC能活化T細胞并刺激低水平的IL-2生成。當Jurkat細胞經PMA和共刺激劑比如植物血凝素PHA處理,IL-2的生成明顯得到加強。PHA本身能夠激活低水平的T細胞活化和IL-2生成,通過非特異性結合到細胞表面受體復合物。T淋巴細胞中,Forskolin激活腺苷酸環化酶會抑制IL-2合成。Forskolin通過間接干擾磷脂酶C活化來發揮這一功能。Forskolin激活腺苷酸環化酶,產生cAMP,從而激活PKA。PKA能磷酸化PLC的絲氨酸殘基,從而間接抑制IL-2生成。

反應步驟:

2.5 mL of Jurkat cells (1×106cells/mL) and 2.5 mL fresh culture medium (RPMI-1640 with 10% fetal calf serum containing 10 mL/L penicillin-streptomycin) were added to 25 cm2culture bottles. The following additions were made in duplicate.

a. Control - no additions

b. 1 μg/mL PHA + 10 ng/mL PMA [Add 10 μL of PHA stock solution (0.5 mg/mL PHA in filter-sterilized PBS) and 5 μL of PMA stock solution (10 μg/mL PMA in DMSO)]

c. 0.2 mM Forskolin + 1 μg/mL PHA + 10 ng/mL PMA [Add 10 μL of PHA stock solution, 5 μL of PMA stock solution, and 10 μL of forskolin stock solution (100 mM forskolin in DMSO)

After mixing well, the bottles were incubated at 37 °C for 24 hours. After centrifugation, the clarified broth was then tested for IL-2 production by ELISA assay. IL-2 production in the test cultures containing 0.2 mM forskolin was inhibited ≥50% compared to the test cultures containing only PHA and PMA. 


產品特性

1)CAS NO:66575-29-9

2)化學名:5-(acetyloxy)-3-ethenyldodecahydro-6,10,10b-trihydroxy-3,4a,7,7,10a-pentamethyl-(3R,4aR,5S,6S, 6aS,10S,10aR,10bS)-1H-naphtho[2,1-b]pyran-1-one

3)同義名:Coleonol洋紫蘇醇;NSC 375489;NSC 357088;L 75-1362B;HL 362;

4)溶解性:溶于乙醇(~15 mg/ml),DMSO(~30 mg/ml),DMF(~30 mg/ml),幾乎不溶于水

5)分子式:C22H34O7

6)分子量:410.50 g/mol

7)純度:>99%

8)外觀:白色結晶性粉末

9)化學結構式:


保存與運輸方法

保存:凍干粉-20oC避光保存,至少3年有效;DMSO儲存液-20oC避光保存至少6個月有效;

運輸:冰袋運輸。


使用方法【源自文獻,僅作參考】

文獻1,Rodriguez G et al.Forskolin-inducible cAMP pathway negatively regulates T-cell proliferation by uncoupling the interleukin-2 receptor complex. J Biol Chem. 2013 Mar 8;288(10):7137-46.PMID: 23341462

體外研究:

細胞類型(Cell type):Human leukemic cell line MT-2 cells

藥物配制(Preparation):Forskolinwas dissolved in DMSO to make stock solution.

實驗方法(Assay):MT-2 cells were seeded into 96-well plates at 5 × 104 cells per well. Cells were then pretreated for1 h with 1% DMSO (vehicle) or Fsk at 1, 5, 10, 25, 50, and 100 μM concentrations. The cells were stimulated with IL-2 as above and cultured for an additional 20 h at 37 °C. Control cells were treated with 1% DMSO for 20 h. During the final 4 h of incubation, the cells were pulsed with [3H]thymidine at a concentration of 0.5 μCi/200 μl. Cells were harvested onto fiberglass filters and analyzed using liquid scintillation counting.Fsk treatment inhibited the proliferation of MT-2 cells in a dose-dependent manner with an IC50 equal to~5 μM Fsk. 

文獻2,Matsumiya W et al. Forskolin modifies retinal vascular development in Mrp4-knockout mice. Invest Ophthalmol Vis Sci. 2012 Dec 7;53(13):8029-35. PMID:23154460

體內研究:

動物模型(Animal Model):C57BL/6J mice and Mrp4-knockout mice

藥物配制(Preparation):Forskolin was dissolved in dimethyl sulfoxide (DMSO) and then diluted.

注射劑量(Dosages):Forskolin was dissolved in DMSO and injected intraperitoneally into neonatal mice at postnatal days 4 (P4) (1.0 μg/50 μL (0.3 mg/kg))and 5 (P5)(1.5 μg/50 μL (0.5 mg/kg)). Mice injected with DMSO served as the controls. The treated mice were euthanized at P6, and their retinas were isolated for whole-mount immunohistochemistry (IHC)

給藥途徑(Administration):Intraperitoneally (i.p.) Injection


訂購信息:進口原料(廣泛應用于全球各大實驗室,純度>99%),質量保證。小規格提供50μl(50mM in DMSO),按照10-50μM的工作濃度來算,可配制50ml-250ml工作液。 

產品名稱

產品編號

規格             

CAS NO.       

價格(元)      

Forskolin (50mM in DMSO) 佛司可林    

MZ2301-50UL        

50μl

66575-29-9

150

Forskolin (50mM in DMSO) 佛司可林

MZ2301-250UL

5×50μl

66575-29-9

520

Forskolin (Powder) 佛司可林

MZ2301-50MG

50mg

66575-29-9

1480

Forskolin (Powder) 佛司可林

MZ2301-100MG

100mg

66575-29-9

2540


注意事項

1)本品不是臨床藥物,只能用于科研用途,不能用于診斷或臨床用途。

2)為了您的安全和健康,請穿實驗服并戴一次性手套操作。


參考文獻

[1] Barber, R. and Goka, T.J. "Adenylate cyclase activity as a function of forskolin concentration." J. Cyclic Nucleotide Protein Phosphor. Res. 10: 23-29 (1985).

[2] Laurenza, A., et al. "Forskolin: a specific stimulator of adenylyl cyclase or a diterpene with multiple sites of action?" Trends Pharm. Sci. 10: 442-447 (1989).

[3] Robbins JD et al. Forskolin carbamates: binding and activation studies with type I adenylyl cyclase. J Med Chem. 1996 Jul 5;39(14):2745-52.

[4] Abe, A. and Karaki, H.J. "Effect of forskolin on cytosolic Ca++ level and contraction in vascular smooth muscle." J. Pharmacol. Exp. Ther. 249: 895-900 (1989).

[5] Insel PA et al. Forskolin as a tool for examining adenylyl cyclase expression, regulation, and G protein signaling. Cell Mol Neurobiol. 2003 Jun;23(3):305-14.

[6] Palmer TD et al. The adult rat hippocampus contains primordial neural stem cells.Mol Cell Neurosci. 1997;8(6):389-404.

[7] Son H et al. Pairing of forskolin and KCl increases differentiation of immortalized hippocampal neurons in a CREB Serine 133 phosphorylation-dependent and extracellular-regulated protein kinase-independent manner. Neurosci Lett. 2001 Jul 27;308(1):37-40.


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