SEC coupled to multiangle light scattering (SEC-MALS) was carried out using a Superdex 200 gel filtration column on an AKTA-FPLC (GE Healthcare) coupled with a DAWN multiple-angle light scattering and an Optilab refractive index system (Wyatt Technology, Santa Barbara, CA). Data for NTD and CTD were collected by injecting 70 and 100 μM protein, respectively, in the FL-N NMR buffer (50 mM sodium phosphate and 150 mM NaCl (pH 6.5)) as well as the NTD NMR buffer (20 mM sodium phosphate and 20 mM NaCl (pH 6.5)). FL-N SEC-MALS data were collected with 20 μM protein in 50 mM sodium phosphate and 150 mM NaCl (pH 7.5). Molar mass and error analysis were determined with ASTRA software package v9, employing a Zimm light scattering model (Wyatt Technology). RNA-binding experiments were carried out with addition 1–1000 RNA to protein to a final ratio of 0.003:1 RNA/protein.
Optilab refractive index system
Manufactured by Wyatt Technology
The Optilab refractive index system is a laboratory instrument designed to measure the refractive index of liquids and solids. It provides accurate and reliable measurements of the refractive index, which is a fundamental physical property of materials. The system is capable of determining the refractive index over a wide range of wavelengths and temperatures.
Automatically generated - may contain errors
Lab products found in correlation
Akta fplc, by GE Healthcare (8 mentions)
Superdex 200 10 300 gl column, by Cytiva (3 mentions)
Astra software package version 8, by Wyatt Technology (3 mentions)
Superdex s200 resin, by GE Healthcare (3 mentions)
Zimm light scattering model, by Wyatt Technology (2 mentions)
Superdex 200, by GE Healthcare (2 mentions)
Dawn multiple angle light scattering, by Wyatt Technology (1 mentions)
9 protocols using optilab refractive index system
1
Multiangle Light Scattering for Protein Analysis
2
Protein Molecular Weight Determination
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Experimental molecular weights were obtained by size exclusion chromatography (SEC) using an AKTA FPLC (GE Healthcare) coupled to a DAWN multi-angle light scattering (MALS) and Optilab refractive index system (Wyatt Technology). Size exclusions were conducted on a Superdex200 10/300 GL column (Cytiva Life Sciences) pre-equilibrated in 50 mM Tris pH 7.4, 150 mM NaCl, 0.5 mM TCEP at room temperature. Protein samples were prepared at 50 μM and injected at a flow rate of 0.8 mL/min. Duplicate datasets were analyzed using ASTRA software package, version 8 (Wyatt Technology).
3
Protein Size Characterization by SEC-MALS
4
Characterizing LC8/syn-4mer Complex
5
Purification and Characterization of BaDV-2 IGR IRES RNA
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In vitro transcribed BaDV-2 IGR IRES RNA was purified by size exclusion chromatography (SEC) with an ÄKTA pure fast protein liquid chromatography (FPLC) (Global Life Science Solutions, New York, NY, USA) as previously described [20 (link)]. RNA-containing fractions were then pooled and concentrated by ethanol precipitation with resuspension in HEPES folding buffer (final concentration: 50 mM HEPES, 150 mM NaCl, 15 mM MgCl2, 3% Glycerol, and at pH 7.4) for SEC-MALS analysis and small-angle X-ray scattering (SAXS) analysis at the concentration of 2 mg/mL. SEC-MALS analysis was performed as previously described with the Optilab Refractive Index System (Wyatt Technology) [20 (link)].
6
SEC-MALS Analysis of Protein Samples
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Size exclusion chromatography in line with multi-angle light scattering (SEC-MALS) was carried out using a SuperdexTM 200 gel filtration column on an AKTA-FPLC (GE Healthcare), a DAWN multiple-angle light scattering, and an Optilab refractive index system (Wyatt Technology). Data for PMDL were collected by injecting 43 μM and 89 μM protein in 150 mM NaCl and 50 mM NaP at pH 7.5 buffer. Data for VP35 65–145 were collected by injecting 100 µM protein in 150 mM NaCl and 50 mM NaP at pH 7.5 buffer. The molar mass and error analysis were determined with ASTRA software package v9, employing a Zimm light scattering model (WYATT Technology, Santa Barbara, CA, USA).
7
Molecular Weight Determination by SEC-MALS
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Experimental molecular weights were obtained by
size exclusion chromatography (SEC) using an AKTA FPLC (GE Healthcare)
coupled to a DAWN multiangle light scattering (MALS) and Optilab refractive
index system (Wyatt Technology). Size exclusions were conducted on
a Superdex 200 10/300 GL column (Cytiva Life Sciences) pre-equilibrated
in 50 mM Tris pH 7.4, 150 mM NaCl, 0.5 mM TCEP at room temperature.
Protein samples were prepared at 50 μM and injected at a flow
rate of 0.8 mL/min. Duplicate data sets were analyzed using the ASTRA
software package, version 8 (Wyatt Technology).
size exclusion chromatography (SEC) using an AKTA FPLC (GE Healthcare)
coupled to a DAWN multiangle light scattering (MALS) and Optilab refractive
index system (Wyatt Technology). Size exclusions were conducted on
a Superdex 200 10/300 GL column (Cytiva Life Sciences) pre-equilibrated
in 50 mM Tris pH 7.4, 150 mM NaCl, 0.5 mM TCEP at room temperature.
Protein samples were prepared at 50 μM and injected at a flow
rate of 0.8 mL/min. Duplicate data sets were analyzed using the ASTRA
software package, version 8 (Wyatt Technology).
8
SEC-MALS analysis of LC8/syn-4mer
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Size-exclusion chromatography (SEC) coupled to a multiangle light scattering (MALS) instrument was performed using an analytical SEC column of Superdex S200 resin (GE Healthcare) on an AKTA-FPLC (GE Healthcare), then routed through a DAWN multiple-angle light scattering and Optilab refractive index system (Wyatt Technology). The column was equilibrated to a buffer of 25 mM tris (pH 7.5), 150 mM NaCl, and 5 mM BME, then injected with 100 μL of LC8/syn-4mer complex in the same buffer at an estimated 2 μM particle concentration (16 μM LC8 + 4 μM syn-4mer, assuming 2:8 binding stoichiometry). We estimated the molar mass using the ASTRA software package, with a Zimm scattering model.
9
SEC-MALS analysis of LC8/syn-4mer
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Size-exclusion chromatography (SEC) coupled to a multiangle light scattering (MALS) instrument was performed using an analytical SEC column of Superdex S200 resin (GE Healthcare) on an AKTA-FPLC (GE Healthcare), then routed through a DAWN multiple-angle light scattering and Optilab refractive index system (Wyatt Technology). The column was equilibrated to a buffer of 25 mM tris (pH 7.5), 150 mM NaCl, and 5 mM BME, then injected with 100 μL of LC8/syn-4mer complex in the same buffer at an estimated 2 μM particle concentration (16 μM LC8 + 4 μM syn-4mer, assuming 2:8 binding stoichiometry). We estimated the molar mass using the ASTRA software package, with a Zimm scattering model.
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