The effects of high pressure and
low temperature on the stability
of two different monoclonal antibodies (MAbs) were examined in this
work. Fluorescence and small-angle neutron scattering were used to
monitor the in situ effects of pressure to infer
shifts in tertiary structure and characterize aggregation prone intermediates.
Partial unfolding was observed for both MAbs, to different extents,
under a range of pressure/temperature conditions. Fourier transform
infrared spectroscopy was also used to monitor ex situ changes in secondary structure. Preservation of native secondary
structure after incubation at elevated pressures and subzero °
C temperatures was independent of the extent of tertiary unfolding
and reversibility. Several combinations of pressure and temperature
were also used to discern the respective contributions of the isolated
Ab fragments (Fab and Fc) to unfolding and aggregation. The fragments
for each antibody showed significantly different partial unfolding
profiles and reversibility. There was not a simple correlation between
stability of the full MAb and either the Fc or Fab fragment stabilities
across all cases, demonstrating a complex relationship to full MAb
unfolding and aggregation behavior. That notwithstanding, the combined
use of spectroscopic and scattering techniques provides insights into
MAb conformational stability and hysteresis in high-pressure, low-temperature
environments.