Theory of Optical Chromatography

Abstract

To evaluate the performance of optical chromatography, a number of equations are theoretically derived using a ray-optics model. These mathematical formalisms are experimentally verified by determining the relationship between the velocity of motion of a polystyrene bead with respect to the intensity of an applied radiation force under the condition where there exists no applied fluid flow. The force is confirmed to be at a maximum at the focal point and to decrease with increasing distance from this position. The radiation force is verified to be proportional to the square of the particle size when the particle diameter is much smaller than the beam diameter. In addition, the radiation force is ascertained to be proportional to the laser power. These results are in excellent agreement with the proposed theoretical model, which is based on ray optics. Furthermore, by analogy with conventional chromatography, fundamental parameters such as retention distance, selectivity, theoretical plate number, and resolution are calculated, and optimum conditions for chromatographic separation are discussed. Based on the results obtained, the dynamic range can be extended by increasing laser power and decreasing flow rate. Peak broadening is primarily caused by variations in laser power and flow rate of the medium for large particles (>1 μm). It is possible, in theory, to distinguish particles whose diameters differ by less than 1% for particles with a diameter larger than 1 μm. Three sizes of polystyrene beads are well separated at a flow rate of 20 μm s^-1 and a laser power of 700 mW. This technique is also applied to the separation of human erythrocytes. Two fractions, one consisting of cells ranging from 1.5 to 2.4 μm in diameter and another consisting of cells ranging from 3.5 to 5.7 μm in diameter, are observed. Optical chromatography is useful for separation and size measurement of particles and biological cells.