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01 Feb 2005
Violet diode lasers pave the way for powerful, cost-effective gene therapy.
A new laser-based technique for introducing foreign genes into cells (a process called transfection) has been demonstrated by scientists in Scotland (Optics Express 13 595). As the process is easy to perform and able to target individual cells it could prove to be an attractive alternative to conventional transfection methods.
Lynn Paterson, Ben Agate and colleagues from the University of St Andrews found that exposing a cell to 0.3 mW of violet light from a diode laser for 40 ms, perforates its membrane allowing the uptake the foreign genes.
The laser beam was focused onto the cell using a x100 microscope objective to get a spot about 1 µm in diameter. The resulting power density of 1200 MW/m2 “punches” a hole into the cell well. The membrane heals itself shortly after the process and the cell does not appear to suffer any long term damage or mutation.
To test the technique the researchers used it to implant an antibiotic resistant gene as well as a green or red fluorescent protein (GFP/RFP) into hamster cells. After inserting the genes the team grew the cells, which appeared to be healthy and multiplied normally. The transfected cells were both fluorescent and resistant to the antibiotic, demonstrating that the gene transfer was successful.
Although laser perforation of cells has been demonstrated before, it has previously required femtosecond Ti:Sapphire lasers, argon-ion lasers or frequency-shifted Nd:YAG lasers which are all far more expensive and much larger than diode lasers.
“In the past the assumption has been that you need a 2-photon process to make it work. People have avoided using blue light as the cell's absorption is very high at that wavelength and they thought that it would damage or kill the cell,” Agate told OLE.
“Our required optical power density with the violet diode laser is six orders of magnitude lower than that required when using cumbersome and expensive femtosecond lasers.”
The team from the university’s schools of physics and biology now plans to investigate the cell puncturing process in more detail. It also wants to explore the idea of simultaneously transfecting several cells using a spatial light modulator to split the laser beam into several paths.
“We believe we have only touched the surface with this technology: the method is simple and inexpensive and could have important bio-medical implications and should find wide use,” said Agate. “Since it also has the potential to assist in the cellular delivery of other bio-molecules, we are now looking at other cell types to see how widely applicable the method proves to be.”
Author
Oliver Graydon is editor of Optics.org and Opto & Laser Europe magazine.
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