Intracellular delivery of antibodies using TAT fusion protein A.
Internalization of antibodies into mammalian cells is a useful method for analyzing and regulating cellular function. In this study, we developed a novel method for the delivery of antibodies into cells using the TAT-fused protein. This fusion protein consists of two functional domains, the protein transduction domain of HIV-1 TAT and the B domain of staphylococcal protein A (SpA), which has an ability to bind to the IgG. The TAT-SpA fusion protein was mixed with fluorescence-labeled rabbit IgG and added to cells. The internalization of antibody was analyzed using confocal microscopy and flow cytometry in living cells. As a result, fluorescence-labeled IgG with the TAT-SpA fusion protein was observed intracellularly. Flow cytometry results demonstrated time course and dose dependence relationships of antibody internalization. These results suggest that the TAT-SpA fusion protein can be a useful reagent for the delivery of antibody into cells.
Mie
Masayasu
M
Department of Biological Information, Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, Yokohama, Japan.
Takahashi
Fumio
F
Funabashi
Hisakage
H
Yanagida
Yasuko
Y
Aizawa
Masuo
M
Kobatake
Eiry
E
D041721:3T3-L1 Cells; D000818:Animals; D000906:Antibodies / Q000737:chemistry*; D015153:Blotting, Western; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D004705:Endocytosis; D005434:Flow Cytometry; D015696:Gene Products, tat / Q000737:chemistry / Q000378:metabolism*; D018014:Gene Transfer Techniques; D015756:Genes, tat / Q000235:genetics*; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000378:metabolism; D051379:Mice; D008957:Models, Genetic; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013205:Staphylococcal Protein A / Q000378:metabolism*; D013997:Time Factors
D000906:Antibodies; D015696:Gene Products, tat; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D013205:Staphylococcal Protein A; D049452:Green Fluorescent Proteins