Which OMNISEC detectors are best for me?
If you are a regular reader of my posts (thanks!), then you know that OMNISEC is a multi-detector GPC/SEC system for the analysis of macromolecules. A fully loaded OMNISEC system contains refractive index (RI), UV-Vis photodiode array (UV-Vis PDA), light scattering, and viscometer detectors. The various detectors and combinations thereof work together to offer different pieces of the characterization puzzle, including molecular weight moments, intrinsic viscosity (IV), and hydrodynamic radius (Rh), to list some examples illustrated in the image below.
One of the advantages of OMNISEC is that the detector configuration in OMNISEC REVEAL is flexible. This is because people use their OMNISEC systems to analyze a wide range of materials for a variety of purposes.
Previously, I’ve described the data that’s available with each detector combination. In this post, I wanted to approach the detector configuration question from a different perspective: what detectors do you need to achieve a specific characterization goal?
In the sections below, I’ll highlight common characterization requests, list the appropriate detector configuration, and share a few notes on each.
Molecular weight & dispersity
Detectors: RI & light scattering
The most common reason to for interest in a GPC/SEC system is to measure molecular weight. Simply put, this is what GPC/SEC does best. This includes molecular weight moments (Mn, Mw, Mz) and dispersity (Mw/Mn). The pairing of light scattering and RI detectors allows the absolute molecular weight to be calculated. With this configuration, the molecular weight values calculated are accurate and there’s no need to generate a calibration curve.
It should be noted that this configuration and the others that include a light scattering detector can utilize the right angle + low angle light scattering detector within OMNISEC REVEAL or the modular SEC-MALS 20 multi-angle light scattering detector. Please visit this link for more info on the two light scattering detector options.
Size, structure, & branching
Detectors: RI, light scattering, & viscometer
The key addition here is the viscometer detector, which allows the shape of the sample molecule in solution to be studied. Intrinsic viscosity measurement requires the presence of a viscometer, but of more frequent interest are the secondary calculations that use IV, such as hydrodynamic radius, Mark-Houwink (MH) parameters & plots (like the ones shown below), and branching calculations.
Data such as IV, Rh, MH parameters, and branching calculations can be determined without a light scattering detector. However, since molecular weight is a required component of those calculations, it is recommended to use a light scattering for absolute molecular weight.
When a multi-angle light scattering detector, such as the SEC-MALS 20, is used for absolute molecular weight, the radius of gyration (Rg) can also be calculated for sufficiently large fractions of the sample.
Compositional analysis
Detectors: RI, UV-Vis PDA, & light scattering
The presence of both RI and UV-Vis PDA detectors allows for sample concentration to be measured from two different perspectives. For example, in the analysis of a PEGylated protein, the RI detector will respond to the concentration of both the PEG and the protein, whereas the UV-Vis PDA detector will only respond to the protein that contains the chromophore. The software can then determine the relative concentration of each component within the sample (shown below).
When a light scattering detector is present, the relative mass of each component can be determined, which in the PEGylated protein case, can help discern how many PEG groups are attached to each protein molecule. Please visit this link for details on compositional analysis.
Aggregate identification
Detectors: RI &/or UV-Vis PDA & light scattering
The light scattering detector is the critical piece here. As detailed in my post on aggregates, the high molecular weight of aggregates produces a strong light scattering response, even if the aggregates are present at low concentrations (which results in low or unobservable RI or UV responses). Like the example shown below, the light scattering detector identifies the aggregate. If there is enough detector response, the RI or UV can potentially quantify the aggregate.
Relative molecular weight & dispersity
Detector: RI (optional UV-Vis PDA also works)
While the accuracy of relative molecular weight values can be hit or miss, using a single detector and a calibration curve is perfect for making comparisons between samples. Or comparing samples to expected results, such as in a QC environment.
The advantage of conventional calibration is that it’s the most economical GPC/SEC setup, featuring the fewest components and simplest calculations. However, using a single detector limits your perspective – you don’t know what you might be missing!
Complete characterization
Detectors: RI, UV-Vis PDA, light scattering, viscometer
A fully loaded OMNISEC system offers everything mentioned above: all of the pieces of characterization data and all of the different analysis options, including the multiple methods to calculate molecular weight. The advantage with this set up is that you can adjust your analysis to meet the needs of your application or samples. It works in research as well as QC, for polymers as well as proteins, and everything in between!
Final thoughts
I hope this post has increased your understanding of how a GPC/SEC system such as OMNISEC can be configured to provide the data you need. If you have any questions, please don’t hesitate to contact us or email me directly at kyle.williams@malvernpanalytical.com.
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