Intracellular Internalization Mechanism of Protein Transfection Reagents

Abstract

A protein transfection reagent is a powerful tool for elucidating a protein function in a cell, and plays an important role in the fields of cell biology and drug discovery. Many researchers have developed protein delivery systems and several systems are commercially available. In this study, we focus on the biological functions of three commercially available protein transfection reagents, Pro-DeliverIN, Xfect, and TurboFect, especially in their internalization routes by HeLa cells. A cellular uptake study using specific endocytosis inhibitors and confocal laser scanning microscope observation revealed that each reagent was internalized into HeLa cells by different mechanism. It is our hope that the results presented here will help in the choice and use of protein transfection reagents for experiments.

Plasmid DNA delivery using fluorescein-labeled arginine-rich peptides

“…The effects of the following endocytosis inhibitors on internalization were examined: amiloride (a specific inhibitor of macropinocytosis); chlorpromazine (a specific inhibitor of clathrin-mediated endocytosis); and nystatin (a specific inhibitor of caveolae-mediated endocytosis). [16][17][18][19] Similar results were obtained from experiments using the R9 peptide alone and R9/pDNA complexes. Cellular uptake was significantly lowered by treatment with amiloride than by the treatment with no additive control, which implied that the internalization of R9 peptides added to peptides alone and peptide/pDNA complexes into Huh-7 cells mainly occurred via macropinocytosis.…”

Molecular logistics using cytocleavable polyrotaxanes for the reactivation of enzymes delivered in living cells

“…4B and 4C). The Xfect internalizes into cells through macropinocytosis or caveolae-mediated endocytosis40. Therefore, Xfect/β-gal is observed as separated from acidic lysosomes because macropinosomes are leaky vesicles, and caveosomes are not involved in the reduction of pH41.…”

Protein Transfection Study Using Multicellular Tumor Spheroids of Human Hepatoma Huh-7 Cells

“…We recently reported the intracellular internalization mechanism of three different commercially available protein transfection reagents, the lipid-based Pro-DeliverIN, peptide-based Xfect, and cationic polymer-based TurboFect [14]. These reagents were internalized into monolayer cultured HeLa cells by different mechanisms, which may be helpful in choosing and using protein transfection reagents for experiments.…”