Automated single-cell sorting system based on optical trapping

Abstract

We provide a basis for automated single-cell sorting based on optical trapping and manipulation using human peripheral blood as a model system. A counterpropagating dual-beam optical-trapping configuration is shown theoretically and experimentally to be preferred due to a greater ability to manipulate cells in three dimensions. Theoretical analysis performed by simulating the propagation of rays through the region containing an erythrocyte (red blood cell) divided into numerous elements confirms experimental results showing that a trapped erythrocyte orients with its longest axis in the direction of propagation of the beam. The single-cell sorting system includes an image-processing system using thresholding, background subtraction, and edge-enhancement algorithms, which allows for the identification of single cells. Erythrocytes have been identified and manipulated into designated volumes using the automated dual-beam trap. Potential applications of automated single-cell sorting, including the incorporation of molecular biology techniques, are discussed.

Orientation of Red Blood Cells and Rouleaux Disaggregation in Interference Laser Fields

“…The RBC's hemoglobins are known to absorb at 514.5 nm [4,5]. The intensity of the incident laser beam and the exposure time of cell suspensions were adjusted in such a way that the action of laser radiation by itself did not cause decomposition of RBC rouleaux and did not produce any pronounced photostress on RBCs.…”

“…The orientation of single RBC in two-beam traps has previously been described experimentally as well as theoretically by computer modeling based on individual beam effect [5]. Instead, we discuss RBC behavior under the influence of gradient dipole forces that result from interaction with the interference laser field.…”

Non-destructive on-chip imaging flow cell-sorting system for on-chip cellomics

“…Efficient and rapid purification of cells has been accomplished with techniques such as fluorescence-activated cell sorting (FACS) (Ogawa et al 1994;Hulett et al 1973;Bonner et al 1972;Herzenberg and Sweet 1976;Assenmacher et al 1995;Johnson et al 2007), magnetic-activated cell separation (MACS) (Imamura et al 1995;Owen and Sykes 1984;Chalmers 2005, 2011), automated single-cell sorting using dual-beam optical trapping (Grover et al 2001), differential adhesion cell sorting (Hsu et al 2008), and disposable microfabricated fluorescence-activated cell sorting (lFACS) (Liu et al 2011;Yoon et al 2011;Azuma et al 2007). In addition, recent advancements in optofluidic flow cytometry, in which optics and microfluidics are used together to create novel functionalities on a small chip, hold great promise for lab-on-a-chip flow cytometry Hatayama et al 1994).…”

Optical tweezers applied to a microfluidic system

“…Another example is the work of Grover et al who showed how optical tweezers can be applied in automated single cell-sorting using peripheral blood as a model system. 12 The erythrocytes were aligned into an upright position along the beam propagation. Trapped cells could then be recognized and sorted out by an imageprocessing system.…”