Construction of a short-wave near infrared spectrofluorometer with diode laser source and charge-coupled device (CCD) detection

摘要

Spectrofluorometers employing xenon arc lamp excitation and photomultiplier tube detectors afford sensitivity over the UV/VIS spectral region for which these instruments were designed, but suffer sensitivity limitations in the short-wave near infrared (NIR) region (800 - 1000 nm) because of their limited source energy and low detector quantum efficiency. To achieve high sensitivity in the NIR region, a 30 mW diode laser source, an imaging spectrograph, and a cryogenically-cooled charge-coupled device (CCD) have been combined in a spectrofluorometer specifically designed for use in the NIR region. The diode laser source incorporates integral source filters, optics, and a beam trap, and utilizes a vertical beam geometry which provides an illuminated volume oriented conveniently for the imaging of fluorescence emissions on the entrance slit of the spectrograph. Data is presented which demonstrates that the temporal and spectral stability of the source is equal or superior to that of an arc lamp for solution- phase fluorometry. In addition to spectral information, the CCD detector provides spatial resolution of fluorescence emissions along the vertical path of the excitation beam. An absolute photometric calibration of the CCD detector, and measurement of its read noise, fixed pattern noise, and linear dynamic range is performed using the photon transfer technique of Janesick, et al. Improvement in the instrument performance by more than six decades is demonstrated by measured LOD of NIR dyes using a commercial SLM 4800 instrument and the new diode laser/CCD arrangement. Origin of the present detection limits is discussed.