Laser-Raman imaging provides a view of microfossils

Using in-situ laser-Raman spectroscopy, researchers at the University of Alabama (UAB; Birmingham, AL) and University of California-Los Angeles (Los Angeles, CA) have developed a noninvasive, nondestructive technique to directly correlate chemical composition with optically discernable morphology in ancient carbonaceous fossils.¹ This technique holds promise for clarifying the nature of tiny fossil-like objects of uncertain origin—whether detected in the Precambrian geologic record, discovered in meteorites, or in samples yet to be brought to Earth from Mars and other planets.

"The purpose of the Raman imagery is to show a composition distribution image that overlays the optical image," explains David Agresti, a professor of physics at UAB. "Paleontologists have been looking for many years for just such a technique."

Laser-Raman spectroscopy was performed with macro, micro, and confocal line-scan imaging options that permitted acquisition of individual point spectra, as well as true Raman images that displayed the distribution of a fossil's molecular components (see figure).

A krypton-ion laser equipped with appropriate optics provided laser wavelengths ranging from blue to infrared, of which 476-, 541-, 568-, 647-, and 752-nm were used, typically at a power of <15 mW over a 1-µm spot. Fossil specimens situated within the uppermost 30 µm or exposed at the upper surfaces of polished petrographic thin-sections were centered in the path of the laser beam projected through a microscope, and rectangular areas of interest were selected for Raman imaging.

Backscattered Raman spectra obtained within the rectangles were collected through the same optical system along micrometer-resolution scan lines, and x-y registrations were automatically recorded to provide a pixel-assigned array of spectral elements, known as "spexels." Two hundred spexels were obtained for each specimen to constitute a virtual image that was then processed into a map-like Raman image, showing the molecular distribution of the fossil structures that produced the Raman spectral bands.

The researchers had expected, in terms of the spectrum, to see a compositional image of graphite, comparable to the wavelength of light. They were pleasantly surprised to find two distinct spectral signatures that represented graphite and a material suggestive of biological origin.

The researchers believe that the results obtained in the initial study demonstrate the ability to apply this technique to in situ analysis of micrometer-sized, organic-walled fossils, as well as providing insight into the molecular makeup and preservation of the carbonaceous matter found in specimens.

Through ongoing studies, the researchers are investigating the extent to which laser-Raman imagery can be expected to provide direct evidence of original biochemical compositions. Since the spatial resolution attained with this technique is limited primarily by diffraction of the excitation laser beam, it is expected that it could also be applied to submicrometer-sized structures.

REFERENCE

1. A. B. Kudryavtsev et al., Proc. National Academy of Sciences 98, 823.