Trace Analysis of Biochemical Substances by Near-Infrared Semiconductor Laser Fluorimetry

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1. Laser fluorimetry is very sensitive and is useful for trace analysis. However, a laser is large and expensive, so that it has not been widely used as a conventional tool for spectrometric analysis. In this study I develop a new spectrometric method using a small and less-expensive semiconductor laser.2. Indocyanine green has an absorption band in a near-infrared region, ant it is found to be quenched by OH radical. By using this reaction scheme peroxidase generating OH radical is sensitively determined. This method allows detection of peroxidase at 10^<-4> U levels. This method is also applied to enzyme immunoassay. Based on sandwich assay, insulin is determined at U/ml levels.3. The oscillating wavelength of the near-infrared semiconductor laser is restricted to 780-900 nm, and then its analytical application is quite limited. When the laser is tightly focused into a non-linear crystal, it generates second harmonic emission (390-450 nm). When a KDP crystal is used, the conversion e … More fficiency was 2.5 x 10^<-6>. It decreases to 1.7 x 10^<-11> when second harmonic emission is generated by the laser medium itself. This emission is used as a light source for an optical fiber sensor and for detection of the end-point in neutralization titration.4. In the near-infrared region only minor species are fluorescent, and most of them are chemically unstable in an aqueous solution. Therefore, it is difficult to apply it to practical analytical applications. A visible semiconductor laser (670 nm), which becomes recently available, is used to overcome this problem. A surfactant is determined after solvent extraction into m-xylene with an ion-pair reagent of Rhodamine 800. It is possible to detect sodium lauryl sulfate at 10^<-7> M levels. For application of visible semiconductor laser fluorimetry to a biochemical molecule, a various procedures are investigated for fluorescence labeling of protein with visible dyes. It is found that oxazine 750 is most suitable as a labeling reagent in this study. Less