Peter M. A. Sloot

Scattering matrix-elements of biological particles measured in a flow through system: theory and practice

Light scattering techniques (including depolarization experiments) applied to biological cells provide a fast nondestructive probe that is very sensitive to small morphological differences. Until now quantitative measurement of these scatter phenomena were only described for particles in suspension. In this paper we discuss the symmetry conditions applicable to the scattering matrices of monodisperse biological cells in a flow cytometer and provide evidence that quantitative measurement of the elements of these scattering matrices is possible in flow through systems. Two fundamental extensions to the theoretical description of conventional scattering experiments are introduced: large cone integration of scattering signals and simultaneous implementation of the localization principle to account for scattering by a sharply focused laser beam. In addition, a specific calibration technique is proposed to account for depolarization effects of the highly specialized optics applied in flow through equipment.

Elastic light scattering from nucleated blood cells: rapid numerical analysis

A model is presented to calculate the light-scatter properties of nucleated blood cells which are mimicked by two concentric spheres. The light-scatter characteristics were derived from the Rayleigh-Debye-Gans approximation and by simultaneous application of field averaging of the internal field and modification of the propagation constant inside the different cellular compartments. It is shown that the results obtained with this simple model are similar to those obtained with more complex and more exact theories. In addition it is demonstrated that simultaneous detection of the light-scatter intensities in the forward-, lateral-, and backscatter directions is required to optimize the detection of different cell types in heterogeneous populations of nucleated blood cells.

Computer-assisted centrifugal elutriation II. Multiparametric statistical analysis

A combination of non-interactive statistical methods is discussed to analyze multiparametric light-scatter data obtained by means of computer-assisted centrifugal elutriation.

Computer-assisted centrifugal elutriation I. Detection system and data acquisition equipment

To optimize cell separations by centrifugal elutriation, we constructed an on-line computer-controlled multiparametric light-scatter system. A bypass sample flow, at the outlet of an elutriation rotor, is hydrofocussed and three scatter parameters of each cell are determined up to a maximum of 15,000 cells/second. The 18-bit representation of the parameter values are cumulatively stored by means of a direct memory access interface. The histogram memory is continuously displayed to provide information on the number and type of cells that are elutriated. A special purpose operating system, implemented on a stand-alone computer configuration, allows a high data-acquisition rate and ample data processing capacity. In addition, a local network driver was constructed to facilitate off-line detailed analysis of the data. The equipment is well suited to monitor the centrigural elutriation process. The flexibility of the system allows an extension of the monitor to computer-controlled elutriation.