Time-resolved angular dependent measurement of triggered light scattering changes in biological suspensions

Abstract

We describe a photometer for time-resolved measurements of small changes in light scattering suited for suspensions of biological material. The time resolution is 35 mus, the amplitude resolution for bovine rod outer segments is typically delta I/I = 5 X 10(-4) at a scattering angle of = 20 degrees. The use of the apparatus is demonstrated by recording the near infrared scattering of bovine rod outer segments after excitation with flashes of green light. Semiconductor detector arrays are arranged centrosymmetrically around a hemispherical cuvette. The optical characteristics of a hemispherical cuvette and the resulting geometry of cuvette and detection are discussed. Calculations of optimal signal transfer and noise of the detectors led to the following arrangement for each scattering angle: pairs of parallel connected photodiodes are fed into several current-to-voltage converters, whose output voltages are summed up by a summing amplifier. For the test of the device so-called N signals of fresh and liquid N2-frozen and thawed ROS samples were measured at four scattering angles simultaneously. A strong angular dependence (difference scattering curve) of the relative light scattering change is seen for fresh ROS which is transformed into a flat curve by freezing and thawing. It is concluded that the competence of the fresh sample to extend the light-induced local events - presumably rhodopsin conformational changes - into the gross-structural range is terminated by freezing.