Hoffmann, Soren VronningBruque, Maria G.Jones, Nykola C.Rodger, AlisonAucamp, JeanDafforn, Tim R.Thomas, Owen R. T.2026-05-222026-05-220175-7571WOS:001721845300001PubMed:41874625ORCID:/0000-0002-7111-3024/work/215200315https://hdl.handle.net/1885/733809304Proper folding of therapeutic monoclonal antibody (mAb) drugs is key to their efficacy and safety. Circular Dichroism (CD) spectroscopy is well suited for investigating the secondary structure of proteins, but the adequacy of the technique for antibody molecules has been severely impacted by the lack of suitable CD reference sets that include spectra of proteins of similar structure. Here we report on the performance of an expanded reference set that includes CD spectra of 14 mAbs, and uses structures derived from homology models. This enhanced reference set is part of a freely available Python-based software package, SSCalcPy-mAb, that includes the new SP-mAb178 reference set for secondary structure analysis using the SELCON3 method. We propose to employ both the secondary structure analysis results and a generated list of the proteins in the reference set that SELCON3 applies to derive the results that ascertain whether an antibody of interest is well-folded or not.Open access funding provided by Aarhus Universitet. European Union’s Horizon 2020 research and innovation programme,101004806,EPSRC Centre for Doctoral Training in Formulation Engineering,EP/L015153/1 This project has received funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement No 101004806 for trans-national access (MOSBRI-2021-049 and MOSBRI-2022-125); the EPSRC Centre for Doctoral Training in Formulation Engineering (Grant No. EP/L015153/1); and Lonza Biologics.7en©2026 The authorsBiotherapeutic proteinsProtein secondary structure analysisQuality controlTherapeutic monoclonal antibodies (mAbs)202610.1007/s00249-026-01828-5105034334816