Deae column

Manufactured by GE Healthcare

Sourced in United States

The DEAE column is a lab equipment product used for ion exchange chromatography. It is designed to separate and purify biomolecules based on their ionic interactions with the column's diethylaminoethyl (DEAE) functional groups. The DEAE column provides a reliable and efficient method for researchers to isolate and concentrate specific proteins, nucleic acids, or other charged molecules from complex sample mixtures.

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Lab products found in correlation

Chitin column, by New England Biolabs (2 mentions) S200 column, by GE Healthcare (2 mentions) Sp column, by GE Healthcare (1 mentions) 5 l fermenter, by Sartorius (1 mentions) Bl21 de3 cells, by Thermo Fisher Scientific (1 mentions) Nh4 2so4, by Carl Roth (1 mentions) 6 m urea, by Merck Group (1 mentions) Sepharose cl 4b column, by Merck Group (1 mentions) Mgcl2, by GE Healthcare (1 mentions) Mgcl2, by Carl Roth (1 mentions) Superdex 200 column, by GE Healthcare (1 mentions)

Close spelling variants of this product

HiTrap DEAE column HiPrep DEAE FF 16/10 column DEAE-Sephadex A-25 (40 mg) column (pyridine acetate form) DEAE Sepharose Fast Flow anion exchange column DEAE Sepharose column DEAE-Sepharose CL-6B column HiPrep DEAE 16/10 column DEAE-cellulose column DEAE-Sepharose anion exchange column HiTrap DEAE FF column

8 protocols using deae column

Purification of Fibrinolytic Venom Components

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Crude venom of NnV (180 mg) was dissolved in 10 mM Tris-HCl buffer (pH 7.8) and centrifuged at 13000×g for 30 min. The venom solution was resolved on DEAE column (GE Healthcare), previously dialyzed against extraction buffer at 4 °C. The column was eluted with gradient concentration of NaCl from 0 to 0.8 M at a flow rate of 1 mL per min. Each peak was tested for fibrinolytic activity and SDS-PAGE.

Bae S.K., Lee H., Heo Y., Pyo M.J., Choudhary I., Han C.H., Yoon W.D., Kang C, & Kim E. (2017). In vitro characterization of jellyfish venom fibrin(ogen)olytic enzymes from Nemopilema nomurai. The Journal of Venomous Animals and Toxins Including Tropical Diseases, 23, 35.

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Purification of Recombinant CYP116B46

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A single transformant of pET46 Ek/LIC expressing wild-type or mutated CYP116B46 gene was grown overnight at 37 °C in LB containing 100 µg mL⁻¹ ampicillin. Ten liters of fresh LB medium with ampicillin (100 µg mL⁻¹) were each inoculated with 200 mL overnight culture and grown to an OD600 of 0.6. The protein expression was induced by 0.4 mM IPTG (isopropyl β-D-1-thiogalactopyranoside) at 16 °C for 18 h. The cells were harvested by centrifugation and then resuspended in 100 mL of lysis buffer containing 20 mM Tris, 150 mM NaCl, and 10 mM imidazole, pH 8.0. Cell disruption was conducted by using a French press (GuangZhou JuNeng Biology and Technology Co. Ltd, Guangzhou, China), and then the lysate was centrifuged at 30,000 × g for 30 min to remove debris. The supernatant was heated at 50 °C for 20 min, and the precipitation was removed by centrifugation at 30,000 × g for 30 min. The extract was loaded onto a Ni-NTA column that was equilibrated by lysis buffer containing 20 mM Tris, 150 mM NaCl, and 10 mM imidazole. The recombinant protein was eluted using a 10–500 mM imidazole gradient. The target protein-containing fractions were collected and further purified by anion-exchange chromatography using a DEAE column (GE Healthcare). Prior to crystallization trials, protein solutions were concentrated to 60 mg mL^-1, and stored at −80 °C.

Zhang L., Xie Z., Liu Z., Zhou S., Ma L., Liu W., Huang J.W., Ko T.P., Li X., Hu Y., Min J., Yu X., Guo R.T, & Chen C.C. (2020). Structural insight into the electron transfer pathway of a self-sufficient P450 monooxygenase. Nature Communications, 11, 2676.

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Purification and PEGylation of Recombinant Proteins

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Example 3

The preparation of mutant ES003 was taken as an example for illustrating the expression and preparation of recombinant ES, ES mutant and Endu mutant. Specifically, the strains for producing ES and its mutants were cultured in a shake flask containing LB medium over night, and then inoculated into a 5 L fermenter (Sartorius). IPTG was added at the appropriate time, and then the bacteria were harvested after about 4 hours (FIG. 2A). The bacteria were resuspended in a buffer solution and deeply broken by a high pressure homogenizer, and the broken bacteria were centrifuged to collect pellets. This process was repeated for three times. Then, DEAE column or Q column (GE Healthcare), and CM column or SP column (GE Healthcare) were used for the elution of proteins with a pH gradient of 4.0 to 10.0. The renatured and un-renatured proteins were purified respectively, so as to obtain the renatured proteins with purity greater than 95% (FIG. 2B, C). The renatured proteins were concentrated and then dialyzed with PBS or NaAc-HAc. The modification of N-terminal of the renatured proteins was performed by using monomethoxy Poly(ethylene glycol)-aldehyde (mPEG-ALD, 20 kDa, Beijing JenKem Technology Co., Ltd.). The modified proteins were purified by using CM column or SP column (GE Healthcare), and eluted with a pH gradient of 4.0 to 10.0 to obtain the target component (FIG. 2D).

US10647968B2. Endostatin mutants with mutations at ATP binding sites (2020-05-12). None [CN], None [CN], None [CN], None [CN], None [CN], None [CN], None [CN]. Inventors: Yongzhang Luo [CN], Peng Liu [CN], Xinan Lu [CN], Yang Chen [CN], Yan Fu [CN], Guodong Chang [CN], Daifu Zhou [CN].

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Synthesis of NP–ZnPc(TAP)4–Ptx Nanoparticles

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β-Mercaptoethanol (2 µL), PBS (750 µL, pH 8), and HSA solution (250 µL, 200 mg/mL) were mixed together. After being stirred for 15 minutes, the solution was diluted using 1 mL PBS (pH 8). Then, ZnPc(TAP)₄ ethanol solution (0.5 mL, 7.6 mM) was added to the solution. After incubation for 5 minutes, Ptx DMSO solution (15.2 mM, 100 µL) was also added to the solution and mixed for another 5 minutes. Finally, another 2 mL ethanol was added dropwise into the solution. After being stirred for 4 hours, PBS (5 mL, pH 8) was added to the reaction solution and the solution dialyzed to PBS (pH 7.4) by dialysis bag with molecular-weight cutoff 8–10 kDa. After the dialysis, the solution was centrifuged at 12,000 rpm for 30 minutes and NP–ZnPc(TAP)₄–Ptx precipitated. NP–ZnPc(TAP)₄–Ptx was resuspended three times for purification. The NP–ZnPc(TAP)₄–Ptx solution was then applied to a diethylaminoethyl anion-exchange (DEAE) column (GE Healthcare UK Ltd, Little Chalfont, UK) for further purification. NP–ZnPc(TAP)₄ or NP–Ptx was prepared by the same method, except for the addition of Ptx or ZnPc(TAP)₄, respectively.

Wang Y., Zheng K., Xuan G., Huang M, & Xue J. (2018). Novel pH-sensitive zinc phthalocyanine assembled with albumin for tumor targeting and treatment. International Journal of Nanomedicine, 13, 7681-7695.

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Purification of Recombinant Human KAT II Enzyme

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The expression plasmid containing human KAT II (hKAT II) as C-terminal intein (chitin binding domain) fusion, in pTXB1 vector, was transformed into BL21 (DE3) cells. Cells were grown in 2YT media at 37°C to an OD₆₀₀ of 0.6, and induced with 300 μM isopropyl ²-D-1-thiogalactopyranoside for 18–20 h at 18°C. The cells were harvested by centrifugation at 2,700 × g for 20 min. The pellets were re-suspended in chitin column buffer, pH 8.5, containing 20 mM Tris, 500 mM NaCl and 1 mM EDTA in 10% glycerol and 5 mM ²-mercaptoethanol. hKAT II was purified by affinity chromatography using a chitin column (New England Biolabs, Ipswich, MA, USA). Intein was cleaved on the column by washing with the chitin column buffer containing 50 mM dithiothreitol. This was followed by ion exchange chromatography using a DEAE column (GE Healthcare Life Sciences, Marlborough, MA, USA). Pure hKAT II was obtained after a final round of purification by size exclusion chromatography, using a S200 column (GE Healthcare Life Sciences). Protein (in 20 mM Tris, 50 mM NaCl and 40 μM pyridoxal-5′-phosphate, pH 8.5) was concentrated to 10.2 mg/ml, flash-frozen in liquid nitrogen and stored at −80°C.

Sathyasaikumar K.V., Tararina M., Wu H.Q., Neale S.A., Weisz F., Salt T.E, & Schwarcz R. (2017). Xanthurenic Acid Formation from 3-Hydroxykynurenine in the Mammalian Brain: Neurochemical Characterization and Physiological Effects. Neuroscience, 367, 85-97.

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Purification of recombinant human KAT II

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The expression plasmid (Genscript, Piscataway, NJ, USA) containing human KAT II (hKAT II) as C-terminal intein (chitin binding domain) fusion, in pTXB1 vector, was transformed into BL21 (DE3) cells (Thermo Fisher Scientific). Cells were grown in 2YT media (Thermo Fisher Scientific) at 37°C until the optical density reached 0.6 (measured at 600 nm), and induced with 300 μM isopropyl 2-D-1-thiogalactopyranoside for 18-20 h at 18°C. The cells were then harvested by centrifugation (2,700 x g, 20 min), and the pellet was re-suspended in chitin column buffer, pH 8.5, containing 20 mM Tris, 500 mM NaCl and 1 mM EDTA in 10% glycerol and 5 mM 2-mercaptoethanol. hKAT II was purified by affinity chromatography using a chitin column (New England Biolabs, Ipswich, MA, USA). Intein was cleaved on the column by washing with the chitin column buffer containing 50 mM dithiothreitol. This was followed by ion exchange chromatography using a DEAE column (GE Healthcare Life Sciences, Marlborough, MA, USA). Pure hKAT II was obtained after a final round of purification by size exclusion chromatography, using a S200 column (GE Healthcare Life Sciences). Protein (in 20 mM Tris, 50 mM NaCl and 40 μM pyridoxal-5′-phosphate, pH 8.5) was concentrated to 10.2 mg/ml, flash-frozen in liquid nitrogen and stored at −80°C.

Ayala T.B., Sathyasaikumar K.V., Uys J.D., Peréz-de-la-Cruz V., Pidugu L.S, & Schwarcz R. (2020). N-Acetylcysteine Inhibits Kynurenine Aminotransferase II. Neuroscience, 444, 160-169.

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In vitro Transcription and Purification of tRNAs

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Human tRNA^Thr and tRNA^Ala was in vitro transcribed as described previously with modification (18 (link)). Template DNA for tRNA transcription was amplified by PCR. After 5-h transcription reaction at 37°C, the reaction was terminated and the product loaded onto a DEAE column (GE healthcare, USA) equilibrated in low salt buffer containing 50 mM MES pH 6.5, 150 mM NaCl, 0.2 mM EDTA to remove proteins and NTPs. The tRNA was eluted with high salt buffer containing 400 mM NaCl. tRNA fractions were pooled and precipitated by ethanol. The purity of tRNA was checked by urea-PAGE. Prior to use in assays, tRNA was incubated at 80°C for 5 min followed by refolding at 60°C in 5 mM MgCl₂ and gradually cooling down to room temperature.

Chen M., Kuhle B., Diedrich J., Liu Z., Moresco J.J., Yates III J.R., Pan T, & Yang X.L. (2020). Cross-editing by a tRNA synthetase allows vertebrates to abundantly express mischargeable tRNA without causing mistranslation. Nucleic Acids Research, 48(12), 6445-6457.

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Purification of Key Cytoskeletal Proteins

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Fibronectin was purified from human plasma by gel filtration and affinity chromatography over a Sepharose CL-4B column (Sigma), followed by a gelatin Sepharose column from GE Healthcare (Munich, Germany). Subsequently, fibronectin was eluted by 6 M urea (Sigma) in PBS and dialyzed against PBS before use.
Actin was isolated from an acetone powder of rabbit skeletal muscle in G-buffer by modifying the protocol of Spudich and Watt. 35 Actin was polymerized by adding 50 mM KCl and 2 mM MgCl 2 (Carl Roth). Subsequently, KCl and MgCl 2 were removed by dialyzation with G-buffer, and the depolymerized actin was purified by gel filtration with a Superdex 200 column (GE Healthcare) and stored in G-buffer. According to the protocol of Margossian and Lowey we also isolated myosin II from rabbit skeletal muscle using centrifugation and salting out. 36 The purified myosin was diluted in D-buffer. a-Actinin was isolated from chicken gizzard following the protocol of Craig et al. 37 After extraction with 1 mM KHCO 3 a-actinin was salted out with (NH 4 ) 2 SO 4 (Carl Roth) and purified with ion exchange chromatography over a DEAE column (GE Healthcare) and gel filtration with a Superdex 200 column. Isolated a-actinin was stored in A-buffer.

Raoufi M., Aslankoohi N., Mollenhauer C., Boehm H., Spatz J.P, & Brüggemann D. (2016). Template-assisted extrusion of biopolymer nanofibers under physiological conditions. Integrative biology : quantitative biosciences from nano to macro, 8(10).

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