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©2021 NanoTemper Technologies, Inc. South San Francisco, CA, USA. All Rights Reserved. 9 APPLICATION NOTE Conclusions There is a complicated matrix of biophysical parameters to evaluate when determining how mutations affect an antibody's stability. Thermal and conformational stability reflect only part of the picture of what characterizes stability parameters. Biologics researchers evaluate a wealth of information to better understand their molecules' early developability profile, and no single experiment tells them everything they need regarding a molecule's attributes. Though there are o en correlations between different stability parameters, they are not strictly linked; for example, a mAb with high thermal stability as reflected by high T m and T on may also demonstrate a high PDI, therefore 55 60 65 70 75 40 50 60 70 80 90 50 55 60 65 70 40 45 50 55 60 50 60 70 80 90 45 55 65 55 65 75 40 60 80 50 60 70 40 50 60 NT.Plex T m1 NT.Plex T on NT.Plex T turb PANTA Scattering IP PANTA T m1 PANTA T on NT.Plex T m1 NT.Plex T turb PANTA T m1 PANTA T on -1.0 -0.8 -0.6 -0.4 -0.2 0.0 0.2 0.4 0.6 0.8 1.0 NT.Plex T m1 NT.Plex T on NT.Plex T turb PANTA Scattering IP PANTA T m1 PANTA T on NT.Plex T m1 NT.Plex T on NT.Plex T turb PANTA Scattering IP PANTA T m1 PANTA T on Correlation Figure 6: Figure 5 shows the high correlation between T m1 , T on , T turb (formerly T agg ), and turbidity IP measured by both the Panta and the NT.Plex. It also shows strong correlation with the scattering IP and cumulant radius change IP from DLS data and the thermal unfolding, as measured by the NT.Plex. Notably, measurements related to the DLS parameters of hydrodynamic radius (r H ) and PDI do not have strong correlation to thermal stability behavior. DLS parameters can be used to supplement thermal unfolding- based data to have additional ranking parameters for antibodies. Figure 5: Clustering of correlations from data obtained from the Prometheus Panta vs Prometheus NT.Plex as plots showing individual data points (top) and heat maps (bottom). There is a strong correlation between parameters obtained on the two different instrument types for T m , T on , Turbidity inflection point (IP), and T turb . DLS parameters PDI and r H do not exhibit strong correlation to the thermal stability data (data not shown).