200 increase 10 300 gl column

Manufactured by GE Healthcare

Sourced in France

The 200 Increase 10/300 GL column is a laboratory equipment product designed for chromatographic separation and purification of biomolecules. It features a column diameter of 10 mm and a bed height of 300 mm, providing a column volume of 23.56 mL. The column is constructed with high-quality materials to ensure reliable and consistent performance.

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Lab products found in correlation

Mini protean tgx precast protein gel, by Bio-Rad (1 mentions) Balb c mice, by Charles River Laboratories (1 mentions) Laemmli sample buffer, by Bio-Rad (1 mentions) Akta pure, by GE Healthcare (1 mentions) Labassay phospholipid kit, by Fujifilm (1 mentions) Bicinchoninic acid bca assay, by Thermo Fisher Scientific (1 mentions) Rprotein a sepharose, by GE Healthcare (1 mentions) Gel filtration calibration kit hmw, by GE Healthcare (1 mentions) Any kd mini protean precast gel, by Bio-Rad (1 mentions) Chemidoc instrument, by Bio-Rad (1 mentions) Akta pure fplc, by GE Healthcare (1 mentions) Kta pure fast protein liquid chromatography system, by GE Healthcare (1 mentions) Vivaspin, by Sartorius (1 mentions)

7 protocols using 200 increase 10 300 gl column

Murine Anti-CD105 Antibody Generation

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The murine monoclonal anti-CD105 antibody (clone K-ro23) was generated by immunization of female BALB/c mice (Charles River, Wilmington, MA) with gamma-irradiated Sp2/0-Ag14 cells (ATCC, Manassas, VA) transfected with human CD105. Immunization was performed intraperitoneally together with CpG oligonucleotides to enhance immunogenicity. Spleen cells were then fused with Sp2/0-AG14 cells according to the protocol established by Kohler and Milstein^{45 (link)}. Supernatants of the resulting hybridoma cells were screened by flow cytometric analysis using CD105⁺ NALM-16 (DMSZ, Braunschweig, Germany) and CD105-transfected Sp2/0 cells, while mock transfected Sp2/0 cells served as negative control. CD105 antibody was purified using protein A affinity chromatography.
For SDS-PAGE analysis, 3 µg of protein was diluted 1:1 with Laemmli sample buffer (Bio-Rad, Hercules, CA) with or without beta-mercaptoethanol (SERVA Electrophoresis, Heidelberg, Germany) as reducing agent. Samples were analysed on a 10% Mini-PROTEAN ® TGX™ Precast Protein Gel (Bio-Rad). Gels were stained with Roti®-Blue colloidal coomassie blue staining solution (Carl Roth, Karlsruhe, Germany). Antibody purity was confirmed by analytical size exclusion chromatography with a 200 Increase 10/300GL column (GE Healthcare, Chicago, IL).

Kauer J., Schwartz K., Tandler C., Hinterleitner C., Roerden M., Jung G., Salih H.R., Heitmann J.S, & Märklin M. (2019). CD105 (Endoglin) as negative prognostic factor in AML. Scientific Reports, 9, 18337.

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Purification and Analysis of Lutein Nanoparticles

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Lutein ND samples were centrifuged at 15,000 × g for 10 min immediately prior to gel permeation chromatography on a Superdex 200 Increase 10/300 GL column fitted to a GE AKTA Pure fast protein liquid chromatography (FPLC) instrument. Four hundred μl aliquots of lutein ND were applied to the column. Samples were eluted with PBS at a flow rate of 0.75 ml/min. Absorbance was continuously monitored at 280 nm, with collection of 2.0 ml fractions. Fractions corresponding to absorbance peaks were pooled and concentrated to ~500 μl by centrifugal filtration (3,000 Da MWCO). The 280 nm absorbance peaks eluting between 8.2 to 10.2 ml and 12.2–14.2 ml, respectively, were analyzed for phospholipid, lutein, and apoA-I content. EYPC content was determined using a LabAssay Phospholipid kit (Wako Pure Chemical Corp., Japan) according to the manufacturer’s instructions. Lutein content was measured spectroscopically and apoA-I content was determined using the bicinchoninic acid (BCA) assay (Thermo-Fisher) with bovine serum albumin as standard.

Moschetti A., Fox C.A., McGowen S, & Ryan R.O. (2022). Lutein nanodisks protect human retinal pigment epithelial cells from UV light-induced damage. Frontiers in nanotechnology, 4, 955022.

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Recombinant Human ACE2 and Spike Protein Production

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To make human ACE2 protein, pcDNA3-sACE2-WT(732)-IgG1⁴³ (link) (Addgene plasmid #154104, gift of Erik Procko) plasmid was transfected into Expi293 cells using PEI at a ratio of 1:3, and the supernatants were collected after five days. hACE2 was purified from the cell supernatant by using rProtein A Sepharose (GE) followed by running through a Superdex 200 Increase 10/300 GL column. For the spike trimer proteins, paH-spike was transfected into Expi293 cells using PEI at a ratio of 1:3, and the supernatants were collected five days later. The spike proteins were purified using Excel resin (Cytiva) according to the manufacturer’s instructions. The molecular weight and purity were checked by running the proteins on SDS-PAGE.

Wang Q., Iketani S., Li Z., Liu L., Guo Y., Huang Y., Bowen A.D., Liu M., Wang M., Yu J., Valdez R., Lauring A.S., Sheng Z., Wang H.H., Gordon A., Liu L, & Ho D.D. (2023). Alarming antibody evasion properties of rising SARS-CoV-2 BQ and XBB subvariants. Cell, 186(2), 279-286.e8.

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Gel Filtration Chromatography of Protein-Ligand Complexes

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A Superdex™ 200 increase 10/300 GL column was run in buffer D (25 mM HEPES, 150 mM NaCl, 5 mM TCEP, pH 7.4) at a flow-rate of 0.5 mL/min and calibrated with the Gel Filtration Calibration Kit HMW (GE Healthcare). A linear fit a plot of K_av vs. log M_r gave R²= 0.99. 30 µM protein was incubated with ligands, e.g., Zn, GTP, and/or GDP, on ice for 1 h, and 100 µL sample injected.

Nairn B.L., Lonergan Z.R., Wang J., Braymer J.J., Zhang Y., Calcutt M.W., Lisher J.P., Gilston B.A., Chazin W.J., de Crécy-Lagard V., Giedroc D.P, & Skaar E.P. (2016). The response of Acinetobacter baumannii to Zinc starvation. Cell host & microbe, 19(6), 826-836.

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Characterization of Cytochrome c Interactions

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Samples including free cytochrome c, CL ND, PC ND, cytochrome c + PC ND and cytochrome c + CL ND, were centrifuged at 15,000 × g for 10 min. A 200 μL aliquot of each sample was diluted with an equivalent volume of 20 mM HEPES, pH 7.4, 150 mM NaCl and loaded onto a Superdex 200 Increase 10/300 GL column fitted to a GE AKTA Pure FPLC instrument. The column was eluted with 20 mM HEPES, pH 7.4, 75 mM NaCl at a flow rate of 0.75 ml/min with absorbance continuously monitored at 280 nm and collection of 2.0 ml fractions. Following chromatography, fractions corresponding to ND or free cytochrome c, respectively, were pooled and concentrated to ~200 μL using a 3 kDa MWCO centrifugal filtration device. Aliquots of each sample were subjected to SDS-PAGE on “Any kD” Mini PROTEAN precast gel (Bio-Rad) and stained with a solution (90% methanol and 10% glacial acetic acid) containing Amido Black (1 g/L) for 30 min. Gels were de-stained in a solution containing 45% water, 45% methanol, 10% glacial acetic acid. Gel images were obtained on a Bio-Rad ChemiDoc instrument. Densitometric analysis of band intensity was performed using NIH ImageJ software v1.52a [22 (link)].

Fox C.A., Lethcoe K, & Ryan R.O. (2021). Calcium-induced release of cytochrome c from cardiolipin nanodisks: implications for apoptosis. Biochimica et biophysica acta. Biomembranes, 1863(12), 183722.

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Purification of Processed Foamate

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Processed foamate or apoLp-III rHDL (400 μl) were applied to a Superdex 200 Increase 10/300 GL column fitted to a GE AKTA Pure FPLC instrument. Samples were eluted with PBS at a flow rate of 0.75 ml/min. Absorbance was continuously monitored at 280 nm with collection of two mL fractions.

Lethcoe K., Fox C.A, & Ryan R.O. (2021). Foam fractionation of a recombinant biosurfactant apolipoprotein. Journal of biotechnology, 343, 25-31.

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Purification and Characterization of FLAG-tagged hTAS1R2

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The FLAG-tagged hTAS1R2 samples that had been eluted from the ANTI-FLAG M2 beads were pooled and concentrated to 0.3–0.5 mg/mL using a 30-kDa MWCO filter column (Vivaspin, Sartorius, Aubagne, France). Then the concentrated FLAG-tagged hTAS1R2 was purified by gel filtration as previously reported^{36 (link)}. The samples were then loaded for gel filtration chromatography (Superdex 200 Increase 10/300GL column) on an Äkta Pure fast protein liquid chromatography system (GE Healthcare, Velizy-Villacoublay, France). The column was equilibrated with 2 column volumes of wash buffer (PBS, 0.1% LMNG, pH 7.3) before the immunopurified FLAG-tagged hTAS1R2 sample was applied. After loading, the column was rinsed with wash buffer at 0.5 mL/min, and the column flow through was monitored by UV absorbance at 280 nm. The molecular masses of the FLAG-tagged hTAS1R2-detergent complexes were estimated by calibrating the column with a gel filtration standard mixture (Sigma-Aldrich). The following standard proteins were used: thyroglobulin (669 kDa), β-amylase (200 kDa), alcohol dehydrogenase (150 kDa), monomeric BSA (66 kDa), carbonic anhydrase (29 kDa), myoglobin (17 kDa) and lysozyme (14.3 kDa). The protein fractions (0.5 mL) were collected using an automated fraction collector. The collected fractions were deposited on SDS-PAGE, stained by Coomassie blue and subjected to immunoblotting analysis.

Belloir C., Brulé M., Tornier L., Neiers F, & Briand L. (2021). Biophysical and functional characterization of the human TAS1R2 sweet taste receptor overexpressed in a HEK293S inducible cell line. Scientific Reports, 11, 22238.

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