A paper1 by researchers at the National Institute of Standards and Technology (NIST) demonstrates that with improved hardware and better signal processing, a powerful form of molecular vibration spectroscopy can quickly deliver detailed molecular maps of the contents of cells without damaging them. Earlier studies have suggested that to be useful, the technique would need power levels too high for cells.
The technique, broadband coherent anti-Stokes Raman scattering (B-CARS), measures the frequencies associated with different modes of vibration of atoms and their bonds in a molecule. The exact mix of these frequencies is an extremely discriminating "fingerprint" for any particular molecule, so Raman spectroscopy has been used as a chemical microscope, able to detail the structure of complex objects by mapping the chemical composition at each point in a three-dimensional space.
In the biosciences, according to NIST chemist Marcus Cicerone, Raman spectroscopy has been used to detect microscopic cellular components such as mitochondria, detect how stem cells differentiate into new forms and distinguish between subtly different cell and tissue types. It can, for example, detect minor differences between various precancerous and cancerous cells, potentially providing valuable medical diagnostic information. Even better, it does this without the need to add fluorescent dyes or other chemical tags to identify specific proteins.
The catch, says Cicerone, is speed. CARS, which uses a pair of lasers to pump up the vibrational states and increase signal, is part of the answer. The current breakthroughs for a broadband CARS instrument developed at NIST since 2004, says Cicerone, gets the same information in 50 ms per pixel.
The new catch is power. The NIST paper describes a combination of improved hardware to gather spectra over a very broad range of wavelengths, and a clever mathematical technique that effectively amplifies the useable signal by examining a portion of signal normally ignored as background interference. The result, says Cicerone, pushes their minimum power level below the damage threshold while retaining the speed of CARS. "We have all the information that you have in a Raman spectrum, but we get it five to 100 times faster," he says, adding that some obvious modifications should push that higher, opening the door to more widespread use of vibrational spectroscopy in both biology and clinical diagnosis.
1. S.H. Parekh et al. Biophysical Journal V: 99 (2010)
Posted by Lee Mather
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