Humanization Strategy Guide

Full replacement optimizes for human protein function accuracy when mouse-human protein homology is high (>85%) and therapeutic objective requires complete human behavior. Domain humanization provides better compromise when discrete epitopes or catalytic regions require human specificity while supporting physiological integration.

Transient expression in cell culture and humanized protein transfection into appropriate cell systems enables preliminary functional characterization. However, in vivo validation requires actual humanized mice confirming expression, localization, and physiological function.

Approximately 60 to 80% of humanized lines show expected functional phenotypes. Unexpected outcomes reflect inefficient expression, inappropriate splicing, intracellular mislocalization, or unanticipated species-specific interaction differences. Multiple independent lines increase likelihood of identifying appropriately humanized founder.

Humanized models often show modified disease phenotypes compared to conventional models due to species-specific pathway interactions. Some humanized models show enhanced or reduced disease severity requiring empirical determination of disease manifestation in humanized context.