©2021 NanoTemper Technologies, Inc. South San Francisco, CA, USA. All Rights Reserved.
5
APPLICATION NOTE
Evaluation of mutations on antibodies to determine how they alter multiple stability parameters
With the establishment of initial T
m
, T
on
, r
H
, and PDI values for the non-mutated antibodies,
it is possible to determine whether the mutant mAbs have significant deviations even
with only modest structural alterations.
Here we have selected two antibody groups and their mutants, representative from each
group of mAbs. In Figure 2A, we see Protein A and its mutants A1-4. Note that overall, the
changes the mutations made to profiles was minimal compared to the differences in the
initial groups, as seen in Figure 1A and 1B. In Figure 2B, we see similar results for Protein
G, where the nanoDSF and turbidity data have broadly similar shapes to the non-mutated
molecule; note that the non-mutated molecule's cumulant radius does not indicate a
large change in size until a much higher temperature than the mutant molecules.
When examining Table 2, it becomes evident that the mutant mAbs did not exhibit
significantly increased stability, either thermal or conformational, compared to the
non-mutated. However, it also exemplifies the differences that structural changes
from single or double mutations can introduce to a molecule. Small changes in the
protein sequence can have significant and measurable effects on the biophysical
characteristics of a mAb.
