On-Line MALS-QELS (Quasi‑Elastic Light Scattering)
While on-line multi-angle light scattering (MALS) is one of the most important techniques for macromolecular characterization, it can be made even more versatile with the addition of a quasielastic light scattering (QELS, a.k.a. dynamic light scattering) module for determination of hydrodynamic radius. QELS can be added to a Wyatt MALS system as a WyattQELS™ module embedded in the MALS instrument, or by connecting the MALS flow cell to a batch DLS instrument such as a DynaPro® NanoStar® or Mobius® via optical fibre. The QELS instruments can be used to determine the hydrodynamic radius, rh , for a variety of samples in a continuous‑flow mode. The combined MALS-QELS system will measure simultaneously rg, rh, and the absolute molar mass.
Wyatt Technology
While on-line multi-angle light scattering (MALS) is one of the most important techniques for macromolecular characterization, it can be made even more versatile with the addition of a quasielastic light scattering (QELS, a.k.a. dynamic light scattering) module for determination of hydrodynamic radius. QELS can be added to a Wyatt MALS system as a WyattQELS™ module embedded in the MALS instrument, or by connecting the MALS flow cell to a batch DLS instrument such as a DynaPro® NanoStar® or Mobius® via optical fibre. The QELS instruments can be used to determine the hydrodynamic radius, rh , for a variety of samples in a continuousâflow mode. The combined MALS-QELS system will measure simultaneously rg, rh, and the absolute molar mass.
Since rg (the root mean square radius) is determined directly from the angular dependence of the scattered light intensity, at least three angles are required to make a reliable measurement. On the other hand, rh is derived from the fluctuations of light scattering intensity because of diffusion of the molecules (aka Brownian motion) and a single measurement angle suffices.
The QELS measurement is performed on-line in the DAWN flow cell. State-of-the-art optical design, high sensitivity, minimal dead volume, and ease-of-use are the hallmarks of the DAWN detectors. These characteristics, among others, enable them to produce superior signalâto-noise, stability, and sensitivity.
An optical fibre receiver is mounted in the read head of the MALS detector at any angular location. The fibre is, in turn, coupled to an avalanche photodiode in an autocorrelator that has been specially modified to accept the signal from the DAWN instrument.
Figure 1: The hydrodynamic radius versus time for BSA and a glycoprotein, obtained from SEC coupled to on-line MALS-QELS detection. The measurements of size (shown here) and molar mass (shown below) can be combined to learn about molecular conformation.
This application note illustrates the results obtained for bovine serum albumin, BSA, and a glycoprotein, which were separated using a size-exclusion chromatograph (SEC). The MALS-QELS detector determined the hydrodynamic radius versus elution time for the two proteins completely independent of the elution time. Figure 1 shows these results with the 90° LS signal superimposed. For these particular samples, rg was below the MALS rh measurement limit of 10 nm. However, simultaneous measurements of MALS and QELS in the integrated system provide complete results for rh and molar mass (even though the small radius cannot be measured by MALS, molar mass measurements are not impacted). Figure 2 shows the molar mass results determined by the DAWN and the RI detector.
Figure 2: Absolute molar mass versus time for BSA and glycoprotein samples superimposed with the signals from 90 LS detector obtained from size-exclusion chromatography with MALS detection. Glycoprotein aggregates with similar molar mass as BSA aggregates elute earlier due to their extended hydrodynamic size. MALS provides accurate molar mass, regardless of elution time.
The MALS-QELS combination allows the simultaneous determination of absolute molar mass, root-mean-square radius (rg from about 10–500 nm), and hydrodynamic radius (rh from about 0.5–300 nm). Conformation results can now be obtained for molecules ranging from 200 g/mol to hundreds of millions of g/mol.
Wyatt Technology
6330 Hollister Avenue, Santa Barbara, California 93117, USA
Tel. +1 (805) 681 9009
Website:www.wyatt.comE-mail:info@wyatt.com
New Study Uses MSPE with GC–MS to Analyze PFCAs in Water
January 20th 2025Scientists from the China University of Sciences combined magnetic solid-phase extraction (MSPE) with gas chromatography–mass spectrometry (GC–MS) to analyze perfluoro carboxylic acids (PFCAs) in different water environments.
The Next Frontier for Mass Spectrometry: Maximizing Ion Utilization
January 20th 2025In this podcast, Daniel DeBord, CTO of MOBILion Systems, describes a new high resolution mass spectrometry approach that promises to increase speed and sensitivity in omics applications. MOBILion recently introduced the PAMAF mode of operation, which stands for parallel accumulation with mobility aligned fragmentation. It substantially increases the fraction of ion used for mass spectrometry analysis by replacing the functionality of the quadrupole with high resolution ion mobility. Listen to learn more about this exciting new development.
A Guide To Finding the Ideal Syringe and Needle
January 20th 2025Hamilton has produced a series of reference guides to assist science professionals in finding the best-suited products and configurations for their applications. The Syringe and Needle Reference Guide provides detailed information on Hamilton Company’s full portfolio of syringes and needles. Everything from cleaning and preventative maintenance to individual part numbers are available for review. It also includes selection charts to help you choose between syringe terminations like cemented needles and luer tips.
The Complexity of Oligonucleotide Separations
January 9th 2025Peter Pellegrinelli, Applications Specialist at Advanced Materials Technology (AMT) explains the complexity of oligonucleotide separations due to the unique chemical properties of these molecules. Issues such as varying length, sequence complexity, and hydrophilic-hydrophobic characteristics make efficient separations difficult. Separation scientists are addressing these challenges by modifying mobile phase compositions, using varying ion-pairing reagents, and exploring alternative separation modes like HILIC and ion-exchange chromatography. Due to these complexities, AMT has introduced the HALO® OLIGO column, which offers high-resolution, fast separations through its innovative Fused-Core® technology and high pH stability. Alongside explaining the new column, Peter looks to the future of these separations and what is next to come.
Oasis or Sand Dune? Isolation of Psychedelic Compounds
January 20th 2025Magic mushrooms, once taboo, have recently experienced a renaissance. This new awakening is partially due to new findings that indicate the effects of psilocybin, and its dephosphorylated cousin psilocin may produce long lasting results for patients who might be struggling with anxiety, depression, alcohol and drug abuse, and post-traumatic stress disorder. Hamilton Company has developed a methodology for the isolation and identification of 5 common psychedelic compounds used in the potential treatment of disease. The PRP-1 HPLC column resin remains stable in the harsh alkaline conditions ideal for better separations.
