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dc.contributor.authorSteiner-Browne, Marina
dc.contributor.authorElcoroaristizabal, Saioa
dc.contributor.authorRyder, Alan G.
dc.identifier.citationSteiner-Browne, Marina, Elcoroaristizabal, Saioa, & Ryder, Alan G. (2019). Using polarized Total Synchronous Fluorescence Spectroscopy (pTSFS) with PARAFAC analysis for characterizing intrinsic protein emission. Chemometrics and Intelligent Laboratory Systems, 194, 103871. doi: 10.1016/j.chemolab.2019.103871en_IE
dc.description.abstractUsing polarized Excitation Emission Matrix (pEEM) spectroscopy to measure the intrinsic emission of proteins offers a potentially useful methodology for a wide variety of potential applications. However, the presence of Rayleigh light scatter causes significant problems when attempting to use Parallel Factor (PARAFAC) and for anisotropy calculations. The use of polarized Total Synchronous Fluorescence Spectroscopy (pTSFS) can minimize Rayleigh scatter and avoid the use of complex data correction methods. Here, we investigated for the first time the use of pTSFS and PARAFAC to analyze the intrinsic emission of an Immunoglobulin (IgG) type protein in its native state. To enable PARAFAC analysis however, TSFS data (which is not trilinear) must first be transformed into an EEM like layout (t-EEM) and this generated a region with no experimentally acquired information (92% of the explained variance, and a much weaker, mostly Tyr related emission with ~3% of the explained variance. The recovery of this Tyr component was only possible because pTSFS measurements were less contaminated by Rayleigh scattering. Changes in Tyr-to-Trp energy transfer rates caused by thermal motion were detected as an increase in Tyr contribution, which could not be resolved with the equivalent pEEM measurements due to light scatter contamination. The increased selectivity, sensitivity, and reproducibility of pTSFS measurements shows that this is a better option than pEEM for fluorescence emission based monitoring of protein structural change or lot-to-lot variance of IgG type proteins.en_IE
dc.description.sponsorshipThis publication has emanated from research supported in part by a research grant from Science Foundation Ireland (SFI) and is co-funded under the European Regional Development Fund under Grand number (14/IA/2282, Advanced Analytics for Biological Therapeutic Manufacture, to AGR). We also thank Agilent Technologies (Mulgrave Victoria, Australia) for the loan of a fluorescence spectrometer.en_IE
dc.relation.ispartofChemometrics And Intelligent Laboratory Systemsen
dc.subjectImmunoglobulin Gen_IE
dc.titleUsing polarized Total Synchronous Fluorescence Spectroscopy (pTSFS) with PARAFAC analysis for characterizing intrinsic protein emissionen_IE
dc.contributor.funderScience Foundation Irelanden_IE
dc.contributor.funderEuropean Regional Development Funden_IE
dc.local.contactAlan Ryder, School Of Chemistry, Room 213, Arts/Science Building, South Cam, Nui Galway. 2943 Email:
dcterms.projectinfo:eu-repo/grantAgreement/SFI/SFI Investigator Programme/14/IA/2282/IE/Advanced Analytics for Biological Therapeutic Manufacture (AA-BTM)./en_IE

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