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Kerafast抗體Anti-Puromycin 3RH11

更(geng)新時間:2022-07-28

簡要描述:

Kerafast抗體Anti-Puromycin [3RH11]:measure rates of global protein synthesis (mRNA translation) in cells or tissue slices incubated with puromycin, or animals treated with puromycin in vivo.

Kerafast抗體Anti-Puromycin [3RH11]


背景介紹:

貨號:EQ0001,嘌(piao)呤(ling)霉素(su)單克隆(long)抗(kang)體(ti)提供了一種(zhong)非放射性方法來(lai)測量與(yu)嘌(piao)呤(ling)霉素(su)孵育的(de)細胞或組織(zhi)切片中的(de)整體(ti)蛋(dan)白質合成(cheng)(mRNA 翻譯)速率,或體(ti)內用嘌(piao)呤(ling)霉素(su)處理(li)的(de)動物。

特色:

* 允(yun)許直接(jie)使用標準免疫化(hua)(hua)學方法對(dui)翻譯進行簡單(dan)的評估(gu)和量化(hua)(hua)

* 傳(chuan)統(tong)脈(mo)沖追蹤方法(fa)的有(you)利替代方法(fa),后者依賴于(yu)放(fang)射性氨基酸(suan)標記

* 與(yu)蛋白質(zhi)印跡和 ELISA 應用兼容

* 使用 Absolute Antibody 的(de)重組平(ping)臺制造(zao),具有(you)來(lai)自(zi)雜交瘤 3RH11 的(de)可變區(qu)(即特異(yi)性)


嘌呤霉素(su)是一(yi)種氨(an)基核苷類(lei)(lei)抗生素(su),來源于(yu)白化鏈霉菌,在核糖(tang)體中發生翻(fan)譯(yi)過程(cheng)中導致(zhi)鏈過早終止(zhi)。該分(fen)(fen)子的(de)(de)(de)一(yi)部分(fen)(fen)類(lei)(lei)似于(yu)氨(an)酰化 tRNA 的(de)(de)(de) 3' 端,使其可用(yong)(yong)于(yu)蛋(dan)白質(zhi)翻(fan)譯(yi)分(fen)(fen)析(xi)。用(yong)(yong)于(yu)監測蛋(dan)白質(zhi)合成的(de)(de)(de)經典(dian)脈(mo)沖追蹤(zong)或淹沒(mei)劑量方(fang)法依賴于(yu)放(fang)射(she)性(xing)蛋(dan)氨(an)酸和(he)(半)氨(an)酸標記的(de)(de)(de)測量。使用(yong)(yong)嘌呤霉素(su)免疫(yi)檢(jian)測進(jin)行(xing)分(fen)(fen)析(xi)是放(fang)射(she)性(xing)氨(an)基酸標記的(de)(de)(de)一(yi)種有利替代(dai)方(fang)法,并(bing)且允許使用(yong)(yong)標準(zhun)免疫(yi)化學(xue)方(fang)法直接評(ping)估/量化翻(fan)譯(yi)


EQ0001,This monoclonal antibody to puromycin provides a non-radioactive method to measure rates of global protein synthesis (mRNA translation) in cells or tissue slices incubated with puromycin, or animals treated with puromycin in vivo.

Highlights:

* Allows for the simple evaluation and quantification of translation directly using standard immunochemical methods

* Advantageous alternative to traditional pulse-chase methods, which rely on radioactive amino acid labeling

* Compatible with Western Blot and ELISA applications

* Manufactured using Absolute Antibody’s Recombinant Platform with variable regions (i.e., specificity) from the hybridoma 3RH11

Puromycin is an aminonucleoside antibiotic, derived from the Streptomyces alboniger bacterium, that causes premature chain termination during translation taking place in the ribosome. Part of the molecule resembles the 3' end of the aminoacylated tRNA, making it useful for protein translation analysis. Classical pulse-chase or flooding dose methods used to monitor protein synthesis rely on the measurement of radioactive methionine and cysteine labels. Analysis using puromycin immunodetection is an advantageous alternative to radioactive amino acid labeling, and allows for the evaluation/quantification of translation directly using standard immunochemical methods


Product Type:Antibody
Name:Anti-Puromycin (3RH11)
Host:Mouse
Isotype:IgG1 kappa
Clonality:Monoclonal
Clone Name:3RH11
Specificity:This antibody recognizes puromycin.
Immunogen:puromycin hydrochloride
Format:Liquid
Purification Method:Protein G purified
Buffer:PBS with 0.02% Proclin 300
Tested Applications:Western blotting (1:1,000), ELISA and Immunofluorescence microscopy.
Storage:+4C (short-term), -20C (long-term); Avoid repeated freeze/thaw cycles.
Shipped:Cold packs


數據(ju)展示:


(A) C2C12 myoblasts were starved of serum and leucine for 2 hr and then IGF-1 and leucine were added to the medium of some of the cells for 45 min. Puromycin (1uM) was added to the medium of some of the cells (lanes 3-6) 30 min before harvest. (B) Quantification of western blot analysis from panel A. (C) In the same study, but in a separate set of culture dishes, cells were incubated with [35S]methionine instead of puromycin and incorporation was measured.



A Western blot was run using the same samples where one set was run on the left side of the gel and the other on the right.  The left side was probed with our original monoclonal anti-puromycin antibody and the other side was probed with the recombinant anti-puromycin antibody, both at 1:1,000 dilution. The secondary antibody was used at the same dilution for both sides and they were both exposed for ~40 sec. The first two samples were from skeletal muscle of mice where the first lane is muscle from the control hindlimb and the other is from a hindlimb that had been immobilized with a cast for three days.  The other 3 lanes are from HEK393T cells: the first lane is from cells incubated in complete medium, the middle lane is from cells incubated for 2 hr in medium lacking glucose and serum, and the last lane is cells incubated for 1.5 hr without glucose or serum and then glucose and serum were returned during the last 30 min.  As expected, puromycin incorporation was lower in the immobilized hindlimb compared to the contralateral control hindlimb, and also in cells deprived of glucose and serum compared to cells in complete medium.  Resupplementation partially restored incorporation.


Kerafast抗體Anti-Puromycin [3RH11]已發表(biao)文獻(xian)

參考文獻:

1. Lee KH, Zhang P, Kim HJ, Mitrea DM, Sarkar M, Freibaum BD, Cika J, Coughlin M, Messing J, Molliex A, Maxwell BA, Kim NC, Temirov J, Moore J, Kolaitis RM, Shaw TI, Bai B, Peng J, Kriwacki RW, Taylor JP. C9orf72 Dipeptide Repeats Impair the Assembly, Dynamics, and Function of Membrane-Less Organelles. Cell. 2016 Oct 20;167(3):774-788.e17.

2. Tom Dieck S, Kochen L, Hanus C, Heumüller M, Bartnik I, Nassim-Assir B, Merk K, Mosler T, Garg S, Bunse S, Tirrell DA, Schuman EM. Direct visualization of newly synthesized target proteins in situ. Nat Methods. 2015 May;12(5):411-4.

3. Gantois I, Khoutorsky A, Popic J, Aguilar-Valles A, Freemantle E, Cao R, Sharma V, Pooters T, Nagpal A, Skalecka A, Truong VT, Wiebe S, Groves IA, Jafarnejad SM, Chapat C, McCullagh EA, Gamache K, Nader K, Lacaille JC, Gkogkas CG, Sonenberg N. Metformin ameliorates core deficits in a mouse model of fragile X syndrome. Nat Med. 2017 Jun;23(6):674-677.

4. Rangaraju V, Lauterbach M, Schuman EM. Spatially Stable Mitochondrial Compartments Fuel Local Translation during Plasticity. Cell. 2019 Jan 10;176(1-2):73-84.e15.

5. Hafner AS, Donlin-Asp PG, Leitch B, Herzog E, Schuman EM. Local protein synthesis is a ubiquitous feature of neuronal pre- and postsynaptic compartments. Science. 2019 May 17;364(6441).

6. H?rnberg H, Pérez-Garci E, Schreiner D, et al. Rescue of oxytocin response and social behaviour in a mouse model of autism. Nature. 2020;584(7820):252-256.


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