Real-time imaging spectrometry for microscope

Significant research attention has focused on the photodynamic diagnosis (PDD) technique, which uses imaging spectrometry to analyze the image and spectra. Up to now, the fluorescence spectrum at only one point in the image can be measured by imaging spectrometry, but it is not enough for diagnosis. The problem is how to obtain the relationship between the exact position and its respective spectrum. In this paper, we demonstrated a novel imaging spectrometry, which can accurately measure the whole surface of tissue instead of a single point. Using this apparatus, we measured the microscope spectrometry image of the hematoxylin-eosin stained tumorous tissue of mice. From the pseudo RGB images of the spectra, tumorous tissues were very clear. In this image, the spectra at arbitrary positions could be taken instantly as well as the transmission and absorption band based on the background spectrum. The spectral intensity distribution images were also obtained at selected wavelengths (for example 425 nm, 505 nm, 535 nm, 600 nm) to get the area distribution of the stained tumorous tissue. In our experiments, we obtained clear spectrometry imaging from weakly illuminated objects. It took 30 seconds to measure a static image of a sample. Moreover, this apparatus is suitable for real-time measurements of living tissue if using laser illumination, a high-speed CCD camera, a fiber, an endoscope and photosensitizer. In the near future, this novel technique can be used in tumor photodynamic diagnosis.