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  • Photoacoustic detection shows its sensitive side

BioPhotonics
Dec 2006
Lauren I. Rugani

Detection of metastasis often proves to be difficult, either because investigation comes too soon in the development of tumor cells or because the cells have begun to disseminate away from the primary tumor site. Current detection techniques include lymph node or bone marrow assays and tracking the spread of stained cells through the lymphatic system. Besides the inaccuracy of such methods, it has been predicted that malignant melanomas can bypass the lymphatic system altogether and instead spread through the circulatory system.

Because tumor cells are estimated to appear in the circulatory system on the order of one cell per 106 normal blood cells, detection techniques must achieve an accurate sensitivity to such low concentrations. To meet this requirement, a team of researchers from the University of Missouri-Columbia has proposed an in vitro photoacoustic detection method, in which pulsed laser energy is absorbed by a chromophore — the region of a cell that is responsible for its color — and acoustic energy propagates as a result of thermoelastic expansion of the cells. Because the signal emitted by melanoma cells is unique, they cannot be mistaken for normal cells.

The detection mechanism employs a frequency-tripled 450-nm Nd:YAG laser from Opotek Inc. of Carlsbad, Calif., that delivers 5-ns pulses to a customized flow cell used for excitation and subsequent acoustic wave detection of a solution containing a tissue sample with a human malignant melanoma cell line. The resulting photoacoustic signals — detected by a polyvinylidene difluoride film attached to the flow cell — are converted to voltage signals.


Researchers customized a flow cell to enable photoacoustic detection of melanoma cells in solution, adding a piezoelectric polyvinylidene difluoride (PVDF) film to record the acoustic signals generated by the cells after they were irradiated with an Nd:YAG laser. Schematic reprinted with permission of Optics Letters.


The investigators tested the sensitivity of the technique by introducing latex microspheres as melanoma tissue phantoms, placing the spheres in saline solution in concentrations ranging from 700 to 7.12 x 106 microspheres per milliliter. Although the detection limit for these trials was 700 spheres per 20 ml of solution, accounting for the area of the excitation path may enable as few as 10 cells to be present to maintain a definitive signal. They found a linear relationship between the concentration of phantom melanoma cells in the solution and the peak excitation value.

They then conducted trials using a concentration of 2.3 x 106 live melanoma cells in the same amount of solution. Irradiation of the sample with 12 mJ of light energy resulted in a peak excitation after 2.5 μs — an expected signal for melanoma. A staining technique applied to the cells revealed that <2 percent of the cells contained melanin. The researchers thus likened melanoma detection to a 4-mV signal for ∋80 melanin-containing cells in the excitation path.

The linear relationship between excitation and cell concentration illustrates the system’s ability to discern different acoustic wave magnitudes and to determine unknown melanoma concentrations by analyzing peak excitation values. The technique can be applied during any stage of metastasis; early detection may help in disease prevention, whereas application in later stages may help analyze the effectiveness of chemotherapeutic treatments.

The researchers plan to advance their studies to clinical trials involving patients with class IV metastasis in an attempt to detect melanoma cells in the bloodstream. Other trials may include attaching secondary molecules or antibodies to enhance detection.

Optics Letters, Oct. 15, 2006, pp. 2998-3000.


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