BIOMEDICAL instrumentation: Spectroscopy provides real-time optical biopsy
A research group at the Institute for Ultrafast Spectroscopy and Lasers, City University of New York (New York, NY), has discovered a fairly noninvasive method to detect cancers in breast tissue using DNA and protein fingerprints derived from a Kubelka-Munk spectral function (KMSF).1 This ratio of absorption to scattering coefficients is obtained from diffuse reflectance.
"The technique still involves the insertion of a needle into the tissue in question," said Robert Alfano, director of the institute, "but does not require significant tissue removal. It stands out from other methods that may fall under the optical-biopsy category because, for perhaps the first time, the results are presented quantitatively in real time. This new approach can help in stereostatic breast biopsy."
Specimens for the experiments included excised benign, adipose, fibroadenoma (the most common neoplasms of the breast), and malignant breast-tissue samples provided by St. Vincent Hospital (Staten Island, NY), Memorial Sloan Kettering Cancer Center (New York, NY), and the National Disease Research Interchange. According to Institute researcher Yuanlong Yang, specimens were not chemically treated or frozen prior to spectroscopic measurements. The sample breakdown was 25 invasive carcinoma, 22 mixed in situ and invasive, 14 fibroadenoma, 44 benign, and 31 adipose specimens.
Although the Kubelka-Munk spectral functions transformed from diffuse reflectance spectrum peaked at 280 nm for a variety of breast tissue studied, the curve for malignant tissue stood out dramatically.
Researchers measured the diffuse reflectance, fluorescence, and excitation spectra in the same location for each sample using an automated dual lamp-based spectrophotometer. Measurements of diffuse reflectance spectra across 250 to 650 nm were produced with the emission and excitation monochromators scanning on the same wavelengths synchronously. The reference standard was b-carotene crystal dissolved in alcohol (0.8 mg/ml), with small particles of titanium oxide added to enhance the scattering signal during diffuse reflectance measurements. According to Alfano, b-carotene was chosen because it can be used to separate fat from other molecular components in breast tissues.
The experiments measured the diffuse reflectance for the tissues and then generated a KMSF from the b-carotene solution, adipose, benign, and malignant tissues (see figure). The resulting plot included several peaks near 550, 480, 280, and 265 nm. "There was no carotene peak at 480 nm in KMSF of benign or malignant breast tissues," said Alfano. "but the feature of the b-carotene appeared in adipose tissue.
"Some features were found in the KMSF curves of malignant, fibroadenoma, and benign breast tissue. For example, the peak near 280 nm, which corresponds to the absorption of proteins, appeared for all tissue, but the amplitude of the malignant tissue was higher than for fibroadenoma and benign tissue. A distinct peak near 265 nm corresponding to the absorption of DNA appeared only for malignant tissue. For fibroadenoma samples, the peak of the KMSF curve near 280 nm varied, with some specimens higher and others lower. Unlike for malignant samples, there was no distinct 265-nm peak. The signal was smaller at 265 nm," said Alfano.
The researchers summarized their findings as follows: "For malignant tissues, the proteins absorption was higher and DNA absorption was observed. For benign tissues, the absorption of proteins was lower and there was smaller DNA absorption. For fibroadenoma tissue, while proteins absorption was sometimes in the range of malignant samples, relatively smaller DNA absorption was found." These findings are especially critical because it is important to distinguish fibroadenomas from malignancies promptly; until now this may not have been possible without a more-invasive biopsy test. [See related story on p. 166 of this issue.]
Paula Noaker Powell
REFERENCE
- Yuanlong Yang et al., "DNA and protein changes in tissues probed by Kubelka-Munk spectral function," to be published in Proc. SPIE, Vol. 3918.