Purified NLGN3 ECD (wild-type or R/C mutant) was concentrated to 1.5 or 1.1 g/L, respectively, and applied onto an ENrich SEC 650 (10 × 300 mm) column (Bio-Rad) pre-equilibrated with 20 mM Tris-HCl buffer (pH 7.5) containing 150 mM NaCl. The MALS data were collected on a DAWN HELEOS 8+ detector (Wyatt Technology) with an RF-20A UV detector (Shimadzu) and analyzed by the program ASTRA (Wyatt Technology).
Dawn heleos 8 detector
Manufactured by Wyatt Technology
Sourced in Japan
The DAWN HELEOS 8+ detector is a multi-angle light scattering device used for the characterization of macromolecules and particles in solution. It measures the intensity of scattered light at multiple angles simultaneously, providing information about the size, molar mass, and conformation of the sample.
Automatically generated - may contain errors
Lab products found in correlation
Rf 20a uv detector, by Shimadzu (3 mentions)
Enrich sec 650, by Bio-Rad (3 mentions)
Superdex 200 10 300 column, by GE Healthcare (1 mentions)
Optilab t rex refractive index detector, by Wyatt Technology (1 mentions)
Astra software, by Wyatt Technology (1 mentions)
Agilent 1100, by Agilent Technologies (1 mentions)
Spd 20a, by Shimadzu (1 mentions)
Rf 10axl, by Shimadzu (1 mentions)
Wyattqels, by Wyatt Technology (1 mentions)
5 protocols using dawn heleos 8 detector
1
Purified NLGN3 ECD Characterization
2
Oligomeric State Determination of XPA Protein
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Multiangle light scattering combined with size exclusion chromatography (SEC-MALS) was used to determine the oligomeric state of purified His-flXPA-StrepII in solution, as previously described65 (link),66 (link). Briefly, 100 μL of His-flXPA-StrepII at concentrations of 65 μM or 80 μM were injected on a Superdex 200 10/300 column (GE Healthcare) pre-equilibrated with XPA MALS buffer (50 mM HEPES, pH 8.0, 200 mM KCl, 5 mM MgCl2, 5 mM DTT, and 10% glycerol) and eluted at a constant flow rate of 0.5 ml/min at room temperature. The molecular mass of the eluting sample was determined using the light scattering signal from a Dawn Heleos 8+ detector and the concentration signal from an Optilab T-rEX refractive index detector (both Wyatt Technologies). Data analysis was carried out with the ASTRA software (version 6.1.5.22, Wyatt Technologies).
3
SEC-MALS Analysis of Protein Samples
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Samples (1 g L−1) were applied onto an ENrich SEC 650 (10 × 300 mm) column (Bio-Rad) with 20 mM Tris-HCl (pH 7.5) buffer containing 150, 300, or 500 mM NaCl. The MALS data were collected by a DAWN HELEOS 8+ detector (Wyatt Technology) with RF-20A UV detector (Shimadzu) and analyzed by the program ASTRA (Wyatt Technology).
4
Size-Exclusion Chromatography of SALM Proteins
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SALM proteins were concentrated to 0.5 g L−1 and applied onto an ENrich SEC 650 (10 × 300 mm) column (Bio-Rad) in 20 mM Tris-HCl (pH 7.4) buffer containing 250 mM NaCl. The MALS data were collected by a DAWN HELEOS 8 + detector (Wyatt Technology) with an RF-20A UV detector (Shimadzu) and analyzed by the program ASTRA (Wyatt Technology).
5
Multi-Detector Characterization of Macromolecules
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The AF4 separation was performed on an Agilent 1100 (Agilent Technologies) combined with a short channel (SC) separation channel linked to Wyatt Eclipse DualTec (Wyatt Technology, Santa Barbara, CA, USA), coupled with a UV-Vis detector SPD-20A (Shimadzu Co., Kyoto, Japan), a fluorescence detector (RF-10AXL, Shimadzu Co.), and a dynamic light scattering detector (WyattQELS, Wyatt Technology) which is an add-on unit connected to the multi-angle light scattering detector (Dawn Heleos 8+ detector, Wyatt Technology). The light scattering detector was equipped with a K5 cell and a GaAs laser operating at 658 nm. The separation channel equipped with a 350 µm spacer (Wyatt Technology), a length of 15.5 cm and a width from 2.1 to 0.1 cm and a regenerated cellulose membrane with a cut-off of 10 kDa from Nadir (Wiesbaden, Germany). The samples were measured at 0.5 s intervals and the UV and MALS signals were simultaneously recorded as fractograms, plots of detector signal intensity versus time. Data acquisition and processing were performed using ASTRA ® ver. 6.1.1.17 (Wyatt Technology).
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