"New microscopic technique reveals for the first time previously unseen molecules in color."

October 22, 2009

A team of Harvard chemists led by X. Sunney Xie has developed a new microscopic technique for seeing, in color, molecules with undetectable fluorescence. The room-temperature technique allows researchers to identify previously unseen molecules in living organisms and offers broad applications in biomedical imaging and research.

The scientists' results are published in the Oct. 22 issue of Nature. Partial funding for the project was provided by the National Science Foundation (NSF).

Fluorescence is a phenomenon in which an electron in a molecule absorbs energy from light and moves to a higher energy level or excited state. The energy of the light is contained in a unit called a photon.

After a very brief stay at the excited state, the electron returns to its previous energy level, or ground state, by emitting a new photon. The energy of the released photon is discharged in wavelengths of detectable visible light lasting only a few billionths of a second.

Many biologically important colored molecules such as hemoglobin--an oxygen-transport protein in red blood cells--absorb light but do not fluoresce. Instead, the electrons in these molecules release their additional but transient energy by converting it to heat.

"Since these molecules do not fluoresce, they have literally been overlooked by modern optical microscopes," Xie said.

To detect non-fluorescent molecules in biological systems, Xie and his team developed a new type of microscopy based on stimulated emission.

Stimulated emission was first described by Albert Einstein in 1917, and was the basis for today's lasers. In a nutshell, it is a process by which an excited-state electron, perturbed by a photon having the correct energy, drops to its ground state producing an additional photon.

Xie's new microscopic technique generates and records a stimulated emission signal by using two carefully timed input and output pulse trains. In the input pulse train, a modulator switches the intensity of the excitation beam on and off at five megahertz, or MHz. The modulation creates a stimulated emission signal at the same frequency. Each train has an incredibly short pulse duration of approximately 200 femtoseconds. A femtosecond is equal to one billionth of one millionth of a second or 10-15 seconds.

The signal produced by the non-fluorescent molecules provides a highly sensitive image of previously "invisible" molecules.

For the full story, please go to:
http://www.nsf.gov/news/news_summ.jsp?cntn_id=115817&WT.mc_id=USNSF_51


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