![]() Samples should be filtered or centrifuged (6000g, 10-30 min) prior to transfer to the clear bottom plate to remove large aggregates and precipitates. Significant aggregates will make accurate size measurements difficult. DLS is very sensitive to small amounts of aggregates.Detergents frequently have micelle size similar to that of proteins and scatter significantly.DLS is compatible with a range of buffers.This can be used to measure protein and modifier masses of integral membrane proteins in detergent micelles and of glycosylated proteins. Combining SEC, MALS and concentration detectors in an SEC-MALS experiment allows for more accurate mass measurements that SEC or MALS alone.īy using two concentration detectors (RI and UV), the molar mass and weight-fraction of a modifier can be determined. Multi-Angle Light static Scattering (MALS) is used to measure light scattering intensity accurately, which is proportional to the weight-averaged mass in solution. Size-exclusion chromatography (SEC) separates molecules based on hydrodynamic volume, but is dependent on similarity to a set of reference standards for accurate mass determination and fails for elongated or sticky proteins. SEC-MALS is used to measure the weight-averaged molar mass of proteins (and other macromolecules) in solution, and can determine oligomeric state and sample polydispersity. The CMI has a SEC-MALS system with a Wyatt Dawn Heleos Multi-Angle Light Scattering (MALS, 18-angle) Detector, with in-line DLS detector and Optilab TrEX refractive index detector. Required for samples with high molecular weight modifiers, such as glycosylation or detergent micelle will require additional analysisĭn/dc measurements of detergent or protein samples Hydrodynamic Radius from DLS (when light scattering signal is sufficient) Includes:Ĭolumn equilibration with your buffer (pH at or below pH 7.5) or with PBSīSA Standard to test column performance and normalize detectors Prior purification by size-exclusion chromatography (SEC) is strongly recommended. Size-exclusion Chromatography with Multi-Angle Light Scattering (SEC-MALS)ĬMI SEC-MALS System: Wyatt Dawn Heleos with in-line DLS, RI UV, Agilent chromatography Standard SEC-MALS Serviceįor proteins and protein complexes between 15 KDa and 1 MDa, without high molecular weight modifiers (modifiers of mass >5% total).
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