Case Studies and Publications

Far-UV CD Analysis During Biotherapeutic Development: A Method Capability Study​

In this study carried out by Novartis, far-UV CD spectroscopy was qualified for the early development of a monoclonal antibody therapeutic. The high sensitivity of the Chirascan V100 enabled low-level detection of an IgG1 mAb in mixture with a structurally highly similar mAb. A demountable short-pathlength cell was used to minimize spectral contributions by formulation buffer excipients. Limit of Detection and Limit of Quantification were established and CD data contributed to a successful biotherapeutic submission to regulatory authorities.​

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Comparison of NIST mAb standard
with known variants

Secondary structure: are spectra
superimposable in far-UV?

Tertiary structure: are small
differences in near-UV significant?

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Differences in secondary structure not significant*
Differences in tertiary structure are significant*

* Tier 2 statistical analysis of data from Chirascan Q100 using 2SD acceptance criteria

Monitor change under stressed conditions
(forced degradation)

Analysis of tertiary structure. Spectra normalized for protein concentration by simultaneous absorbance measurements.
Chirascan™ Q100, n=6

Monitor change under stressed conditions (forced degradation)

High sensitivity CD analysis of IgG1 samples subjected to a range of degradation conditions revealed minor differences in tertiary structure when compared to a control sample.

Weighted spectral difference method: Dinh, Nikita et al. Anal. Biochem. 464 (2014): 60-62

Quantifiable assessment of changes in tertiary structure

Statistical analysis enabled objective confirmation of spectral results. Degradation conditions have affected local environment of aromatic side chains (no changes were detected in secondary structure – results not shown).

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