Biotechnol. Prog.,
Copyright © 1997 American Chemical Society and American Institute of Chemical Engineers
Selective Precipitation of Antibody with Ligand-Modified Phospholipids: Effect of Lipid Chain Length
Accepted
The selective precipitation from aqueous solutions of goat polyclonal
anti-biotin antibody (pABA)
by complexation with ligand-modified
phospholipids (LMPs) is described. In this study, the
effect of varying the acyl chain length of the LMP
from six to 18 carbon atoms on the rate and yield of precipitation is detailed.
As the acyl chain length increases, the hydrophobic
driving force for interaction of ligand-bound
antibody molecules also increases, resulting in a larger yield of precipitated
antibody. The rate of selective precipitation, however, is observed to pass
through a sharp maximum at an acyl chain length of
10-12 carbon atoms. In the range of target antibody and LMP concentrations
studied (1-10 
M), the
maximum rates of precipitation are observed for those LMPs
in sufficiently low concentrations in aqueous solution to be below their
critical micelle concentration (CMC). The longer chain length LMPs (12-18 carbon atoms at concentrations of 5-10 M) gave
considerably slower rates of precipitation and were all observed to be micellar solutions. The yield of target antibody as a
percentage of antibody precipitated was not observed to pass through a maximum,
rather all LMPs with acyl
chain lengths longer than 12 carbon atoms were observed to give the maximum
yield. Thus the optimal structure of an LMP for precipitation of a target
antibody corresponds to the maximum chain length (10 carbon atoms) at a
concentration level (5-10 M) which
still falls below its CMC. The kinetics of precipitation, as monitored by
measuring turbidity, are well modelled by a theory
which combines the Mie theory of light scattering
with the Smoluchowski theory for the kinetics of
precipitation. The maximum rate constants corresponding to Smoluchowski
kinetics for precipitating pABA
were approximately 25 000-30 000 M-1 s-1, while the
maximum yields were 65-70%. The molecular picture which emerges is one in which
the optimal rate is obtained by maximizing hydrophobic driving force for
interaction of LMP acyl chains while still maintaining
a submicellar state of aggregation.