A sensitive dual wavelength microspectrophotometer for the measurement of tissue fluorescence and reflectance

The construction of a microscope photometer using prefabricated elements is described. To illuminate the tissue, a Leitz Ultropac is applied. To enlarge the wavelength range, its illuminating glass lens is replaced by an Acryl glass zonal lens. Two separate light channels with separate lamps, monochromators and photomultipliers allow the measurement of fluorescence excitation and emission spectra as well as of reflection spectra. By chopping the light, light pulses and dark currents are measured 8.33 times a second. By an integration circuit the signal-to-noise ratio for small signals is improved. The instrument allows us to detect the increase of 4 ng/ml NADH (solution of pH 7.38) in an area of 0.2 mm2.