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.
Zimm light scattering model
Manufactured by Wyatt Technology
Sourced in United States
The Zimm light scattering model is a mathematical model used to analyze the results of static light scattering experiments. It provides a way to determine the molar mass, radius of gyration, and second virial coefficient of macromolecules in solution. The model is based on the Rayleigh-Gans-Debye scattering theory and is commonly used in the characterization of polymers, proteins, and other large molecules.
Automatically generated - may contain errors
Lab products found in correlation
Optilab refractive index system, by Wyatt Technology (2 mentions)
Akta fplc, by GE Healthcare (2 mentions)
Superdex 200, by GE Healthcare (2 mentions)
Dawn mals detector, by Wyatt Technology (1 mentions)
Optilab refractive index detector, by Wyatt Technology (1 mentions)
Superdex 200, by Cytiva (1 mentions)
Astra 6, by Wyatt Technology (1 mentions)
Kta basic system, by GE Healthcare (1 mentions)
Superdex 200 increase 10 300 gl, by GE Healthcare (1 mentions)
Superdex 200 10 300 gl, by GE Healthcare (1 mentions)
Dawn heleos, by Wyatt Technology (1 mentions)
Dawn multiple angle light scattering, by Wyatt Technology (1 mentions)
4 protocols using zimm light scattering model
1
Multiangle Light Scattering for Protein Analysis
2
SEC-MALS Analysis of Protein Molar Mass
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SEC-MALS was carried out using a Superdex 200 gel filtration column on an AKTA fast liquid chromatography system (Cytiva Life Sciences) coupled with a DAWN MALS detector and an Optilab refractive index detector (Wyatt Technology, Santa Barbara, CA). Data for IC1-260 was collected for protein samples at a concentration of 200 µM protein in a buffer composed of 50-mM sodium phosphate (pH 7.5), 50-mM sodium chloride, and 1-mM sodium azide. Data for ICFL was collected for protein samples at a concentration of 30 µM in a buffer composed of 25-mM tris(hydroxymethyl)aminomethane hydrochloride (pH 7.4), 150-mM KCl, 5-mM β-mercaptoethanol, and 1-mM sodium azide. Molar mass and error analysis were determined using ASTRA v9, employing a Zimm light scattering model (Wyatt Technology).
3
Nup82 Complex Molecular Mass Analysis
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Affinity-purified Nup82 complexes were analyzed by SEC equilibrated in a buffer without NP-40 (20 mM Hepes, pH 7.5, 150 mM NaCl, 50 mM K(OAc), 2.5 mM Mg(OAc)2, and 5% glycerol). They were separated on a Superdex 200 10/300 GL or Superdex 200 Increase 10/300 GL column attached to the ÅKTA Basic system (GE Healthcare).
The chromatography system was connected in series with an eight-angle light scattering detector (DAWN HELEOS; Wyatt Technology Corp.) with a light-scattering refractometer for differential refractive index detection (SEC-3010; WGE Dr Bures) was used to determine the molecular mass of complexes. Data were collected at a flow rate of 0.5 ml/min at 4°C. Measurement analysis were performed using the ASTRA 6.1 software and Zimm light-scattering model (Wyatt Technology Corp.), yielding the molecular mass and mass distribution (polydispersity) of the sample.
The chromatography system was connected in series with an eight-angle light scattering detector (DAWN HELEOS; Wyatt Technology Corp.) with a light-scattering refractometer for differential refractive index detection (SEC-3010; WGE Dr Bures) was used to determine the molecular mass of complexes. Data were collected at a flow rate of 0.5 ml/min at 4°C. Measurement analysis were performed using the ASTRA 6.1 software and Zimm light-scattering model (Wyatt Technology Corp.), yielding the molecular mass and mass distribution (polydispersity) of the sample.
4
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).
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