Light scattering is a powerful tool for the characterization of biopharmaceutical products. The two major categories of light scattering techniques include static [Examples: static light scattering (SLS) and multi-angle light scattering (MALS)] and dynamic light scattering (DLS). In the biopharmaceutical industry, these techniques are frequently used for the determination of molecular weight and size of the molecule during product development, downstream processing, product characterization, quality control, and biosimilarity studies.
MALS is a robust and sensitive technique that measures the light scattered by the sample at different angles and calculates the molecular weight using first principles. The MALS detectors available in the market consist of 3-angle and up to 18-angle detectors. Size exclusion chromatography is commonly coupled with a MALS detector to separate monomers from aggregates, fragments, and other impurities. The set-up of SEC-MALS consists of a SEC-UV detector (HPLC) connected to MALS and a differential refractive index (RI) detector. The SEC separates molecules in the solution based on hydrodynamic volume. MALS detector measures the light scattered by the sample. The RI detector determines protein concentration by measuring the difference in refractive indices between the eluate and solvent. Along with SEC, other chromatographic techniques such as reverse-phase (RP), ion-exchange chromatography (IEX), and hydrophobic interaction chromatography were coupled with MALS detector for protein characterization[1][2][3]. Field flow fractionation (FFF) coupled with MALS is also an emerging technology that is used as an alternative technique to SEC and helps to overcome the limitations of SEC[4].
The applications of MALS consist of the determination of molecular weight, conformation, conjugation ratio, radius of gyration, and second virial coefficient (A2) [5][6]. SEC-MALS serves as a powerful analytical technique in exploring the stability and aggregation profile of the product. It is also used to characterize the degree of PEGylation in therapeutic proteins[7]. SEC-MALS along with online dynamic light scattering measures hydrodynamic radius.
MALS and FFF professionals from Wyatt, Spinco Biotech Pvt Ltd, and other experts from the analytical field and biopharmaceutical industry will be attending Biosimilar Workshop 2023 at Goa.
If you wish to learn, explore more, and join us at Biosimilar Workshop 2023, Goa-
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References:
[1] H. Amartely, O. Avraham, A. Friedler, O. Livnah, and M. Lebendiker, “Coupling Multi-Angle Light Scattering to Ion Exchange chromatography (IEX-MALS) for protein characterization,” Sci. Rep., vol. 8, no. 1, Art. no. 1, May 2018, DOI: 10.1038/s41598–018–25246–6.
[2] L. Gentiluomo, V. Schneider, D. Roessner, and W. Frieß, “Coupling Multi-Angle Light Scattering to Reverse-Phase Ultra-High-Pressure Chromatography (RP-UPLC-MALS) for the characterization monoclonal antibodies,” Sci. Rep., vol. 9, no. 1, Art. no. 1, Oct. 2019, DOI: 10.1038/s41598–019–51233–6.
[3] B. A. Patel et al., “Multi-angle light scattering as a process analytical technology measuring real-time molecular weight for downstream process control,” mAbs, vol. 10, no. 7, pp. 945–950, Sep. 2018, DOI: 10.1080/19420862.2018.1505178.
[4] “Multi-Angle Light Scattering with Field-Flow Fractionation.” https://www.wyatt.com/solutions/techniques/fff-mals-characterization-of-nanoparticles-colloids-macromolecules.html (accessed Dec. 09, 2022).
[5] “SEC-MALS,” Wyatt Technology. https://www.wyatt.com/solutions/techniques/sec-mals-molar-mass-size-multi-angle-light-scattering.html (accessed Dec. 09, 2022).
[6] Y. Ma et al., “Determination of the second virial coefficient of bovine serum albumin under varying pH and ionic strength by composition-gradient multi-angle static light scattering,” J. Biol. Phys., vol. 41, no. 1, pp. 85–97, Jan. 2015, DOI: 10.1007/s10867–014–9367–7.
[7] J. Moore and E. Cerasoli, “Particle Light Scattering Methods and Applications,” in Encyclopedia of Spectroscopy and Spectrometry (Third Edition), J. C. Lindon, G. E. Tranter, and D. W. Koppenaal, Eds. Oxford: Academic Press, 2017, pp. 543–553. DOI: 10.1016/B978–0–12–803224–4.00040–6.
