Effects of employment of distinct strategies to capture antibody on antibody delivery into cultured cells.

Abstract

The characteristics of antibody delivery into cultured HeLa cells were examined using two delivery systems. Both systems used a cell-penetrating peptide as a tool for intrusion of an antibody into the cells, but either a "protein A derivative" or "hydrophobic motif" was employed to capture the antibody. When we examined the uptake of the Alexa Fluor-labeled antibody by the use of these two systems, both systems were found to effectively deliver the antibody into the cultured cells. However, when we compared the amount of antibody delivered by these systems with the amount of transferrin uptake, the former was 10 times smaller than the latter. The lower efficiency of antibody delivery than transferrin uptake seemed to be attributable to the involvement of the antibody delivery reagent, which failed to catch the antibody molecule. This interpretation was validated by an experiment using a larger amount of antibody, and the amount of antibody delivered by the "protein A derivative" system under this condition was determined to be 13ng proteins/10(5)cells. The antibody delivery achieved by the "protein A derivative" or "hydrophobic motif" showed two differences, i.e., a difference in intracellular distribution of the delivered antibody molecules and a difference in the fluorescence spectrum observed with cellular lysates. Possible reasons for these differences between the two delivery systems are discussed.