Scanning Microspectrophotometer Using Flow Cytometry

This instrument was designed automatically to detect and classify cells as normal, pre-cancer and cancerous. The instrument provides automated detection and identification of unstained exfoliated cells and live bacteria pathogens in real time. In addition to screening for early lung cancer, it could provide constant monitoring, in real time, of the nation’s water supply and provide an alarm upon the detection of suspicious bacterial pathogens. This instrument is an upgraded and patentable digital version of a prior successful cytological screening instrument. (US Patent # 3,826,899).

PastedGraphic-1For reliable automation of cellular or bacteria specimens, the introduction of dyes to a specimen does not yield repeatable results. Dye uptake is not stoichiometric (a defined ratio among chemicals) – therefore is not reproducible. As a result, even carefully controlled or automated staining techniques seldom yield quantitatively reproducible results. Fluorescent dyes may compete for absorption sites with non-fluorescent impurities, debris and overlap, which further complicate automation techniques. Our use of spectral absorption and mensuration (geometry applied to the computation of lengths, areas, or volumes from given dimensions or angles), using our algorithms, is cost effective, can be done in real time and will have a higher confidence level of accuracy.

The Scanning Microspectrophotometer, uses an ultra violet (UV) source. This UV light beam produces a “raster” scan (a scan pattern in which an area is scanned from side to side in lines from top to bottom) by horizontal and vertical mirror motion. This is created by two sets of front surfaced oscillating mirrors, one mirror at 60 hertz and the other at 15.7 kilohertz. The generated UV raster is directed through a 100X quartz objective lens. This produces a raster spot size in the order of 0.5 micron.

Positive pressure is applied to a fluid specimen to produce laminar flow in a quartz capillary tube. The laminar flow tends to maintain the cells and/or bacteria in the center of the capillary to maintain good focus. The specimen material from the capillary can be collected onto a slide if manual review is required.

As the cells and/or bacteria flow past the objective lens raster scan, the detectors in the spectrometer develop a UV absorbance signal with each slice (63.5 micro seconds) of the UV beam. Since we know the speed of the raster scan, we can do mensuration of each bacteria. The spectrometer output is fed into a computer, which has installed pattern recognition algorithms to identify cells and/or bacteria in real time by their size, shape, spectral absorption and other parameters.

The commercialization of this technology, using sputum cytology for the early detection of lung cancer, also has great commercial potential especially in the real time monitoring of the nation’s water supplies for pathogen detection where time is of the essence to protect the population.