A practical device for pinpoint delivery of molecules into multiple neurons in culture.
We have developed a device for pinpoint delivery of chemicals, proteins, and nucleic acids into cultured cells. The principle underlying the technique is the flow of molecules from the culture medium into cells through a rupture in the plasma membrane made by a needle puncture. DNA transfection is achieved by stabbing the needle tip into the nucleus. The CellBee device can be attached to any inverted microscope, and molecular delivery can be coupled with conventional live cell imaging. Because the position of the needle relative to the targeted cultured cells is computer-controlled, efficient delivery of molecules such as rhodamine into as many as 100 HeLa cells can be completed in 10 min. Moreover, specific target cells within a single dish can be transfected with multiple DNA constructs by simple changes of culture medium containing different plasmids. In addition, the nano-sized needle tip enables gentle molecular delivery, minimizing cell damage. This method permits DNA transfection into specific hippocampal neurons without disturbing neuronal circuitry established in culture.
Hara
Chikako
C
Laboratory for Cell Function Dynamics, Advanced Technology Development Group, Brain Science Institute, RIKEN, 2-1 Hirosawa, Wako-city, Saitama, 351-0198, Japan.
Tateyama
Kiyohiko
K
Akamatsu
Naoki
N
Imabayashi
Hiroyuki
H
Karaki
Koichi
K
Nomura
Nobuo
N
Okano
Hideyuki
H
Miyawaki
Atsushi
A
D000818:Animals; D018929:Cell Culture Techniques; D002478:Cells, Cultured; D004247:DNA / Q000235:genetics / Q000494:pharmacology*; D018274:Electroporation / Q000295:instrumentation / Q000379:methods*; D005456:Fluorescent Dyes / Q000494:pharmacology; D018014:Gene Transfer Techniques / Q000295:instrumentation*; D005822:Genetic Vectors / Q000235:genetics / Q000494:pharmacology; D006367:HeLa Cells; D006624:Hippocampus / Q000187:drug effects / Q000648:ultrastructure; D006801:Humans; D008845:Microinjections / Q000295:instrumentation / Q000379:methods*; D008855:Microscopy, Electron, Scanning; D009339:Needles / Q000592:standards / Q000639:trends; D009474:Neurons / Q000187:drug effects* / Q000648:ultrastructure; D010957:Plasmids / Q000235:genetics / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D012235:Rhodamines / Q000494:pharmacology; D012984:Software; D013997:Time Factors; D014162:Transfection / Q000295:instrumentation / Q000379:methods*; D014584:User-Computer Interface
D005456:Fluorescent Dyes; D012235:Rhodamines; D004247:DNA