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Cytometry - Rapid Label-free Raman Spectroscopy

Current methods that are used for detection, differentiation, and quantitation of individual cells are often time consuming, nonspecific, invasive, and destructive. Discrimination of neoplastic cells from normal cells relies on the use of histochemical stains, exogeneous labeling of surface markers, light microscopy, and cytogenetics. We are developing a rapid technique that can accurately and non-destructively identify and sort neoplastic (e.g. leukemic) cells from healthy cells for the diagnosis and potential treatment of cancer.

A team of spectroscopists and microscopists from UCD and LLNL with clinical oncologists at UCDMC have combined micro-Raman spectroscopy with laser-trapping to obtain highly specific and reproducible molecular signatures of individual cells for the purpose of cell classification, identification, and sorting while leaving the cells alive and intact.

Small particles including single living cells, due to the way they scatter light, can be trapped in the focus of a laser beam that is directed through an optical microscope. In addition to the trapping, some of the laser light excites vibrations in the biomolecules that make up the cell. This is inelastic scattering of light is called Raman scattering. The purpose of the laser trap is to suspend the cells away from surfaces that can typically produce a large background and to hold the sample within the focus of the microscope long enough to obtain a spectrum.

By analyzing the amount of energy lost by the Raman scattered light, a spectrum can be produced that when interpreted, gives a biochemical inventory of the cell contents. Using this technique, we are able to distinguish different types of cells, different states of the same cell, and normal vs. diseased cells.

Multiplexed LTRS image

Figure 1.   Top: Multifocal laser tweezers are used to trap and align suspension particles adjacent to each other. 
Middle: Raman spectra of seven red blood cells acquired simultaneously using the multifocal laser tweezers Raman spectroscopy system.  Bottom: The spectra of a red blood cell on its own and one with a polymer bead in close proximity demonstrate that crosstalk between channels is not an issue in the multiplexed configuration.

Project PI:
James Chan, PhD
eFellow: Lingbo Kong, PhD
Keaton Raphael Memorial Partner:  Teresa Hofhenke

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