Mark M. Wang

Microfluidic sorting of mammalian cells by optical force switching.

Microfluidic-based devices have allowed miniaturization and increased parallelism of many common functions in biological assays; however, development of a practical technology for microfluidic-based fluorescence-activated cell sorting has proved challenging. Although a variety of different physical on-chip switch mechanisms have been proposed, none has satisfied simultaneously the requirements of high throughput, purity, and recovery of live, unstressed mammalian cells. Here we show that optical forces can be used for the rapid (2–4 ms), active control of cell routing on a microfluidic chip. Optical switch controls reduce the complexity of the chip and simplify connectivity. Using all-optical switching, we have implemented a fluorescence-activated microfluidic cell sorter and evaluated its performance on live, stably transfected HeLa cells expressing a fused histone–green fluorescent protein. Recovered populations were verified to be both viable and unstressed by evaluation of the transcriptional expression of two genes, HSPA6 and FOS, known indicators of cellular stress.

Optical Manipulation of Objects and Biological Cells in Microfluidic Devices

In this paper, we review optical techniques used for micro-manipulation of small particles and cells in microfluidic devices. These techniques are based on the objects interaction with focused laser light (consequential forces of scattering and gradient). Inorganic objects including polystyrene spheres and organic objects including biological cells were manipulated and switched in and between fluidic channels using these forces that can typically be generated by vertical cavity surface emitting laser (VCSEL) arrays, with only a few mW optical powers. T-, Y-, and multi-layered X fluidic channel devices were fabricated by polydimethylsiloxane (PDMS) elastomer molding of channel structures over photolithographically defined patterns using a thick negative photoresist. We have also shown that this optical manipulation technique can be extended to smaller multiple objects by using an optically trapped particle as a handle, or an “optical handle”. Ultimately, optical manipulation of small particles and biological cells could have applications in biomedical devices for drug discovery, cytometry and cell biology research.

Parallel transport of biological cells using individually addressable VCSEL arrays as optical tweezers

We have demonstrated the use of vertical cavity surface emitting lasers (VCSELs) for optical trapping and active manipulation of live biological cells and microspheres. We have experimentally verified that the Laguerre‐Gaussian laser mode output from the VCSEL functions just as well as the traditional Gaussian fundamental laser mode for optically trapping biological cells and may be preferable since the highest intensity of the Laguerre‐Gaussian mode is located at the outer ring of the optical aperture, which allows for stronger optical confinement to be obtained for a lower total power. Another advantage that VCSELs have over conventional gas and diode lasers is their ability to be manufactured in an array form. Using a 2 � 2 array of VCSELs, the simultaneous and independent transport of four human red blood cells is demonstrated indicating that much larger two-dimensional VCSEL arrays can be used as individually addressable optical tweezers in biological chips and systems. This parallel transport capability will have a significant impact in currently developing biochip array and assay technologies through the facilitation of the selection, relocation, and precision placement of cells. # 2002 Elsevier Science B.V. All rights reserved.

Experimental characterization of a two-photon memory

We demonstrate the recording of 100 planes of digital images in a page-oriented two-photon memory and characterize the images in terms of signal-to-noise ratio and bit error rate. Possible error sources in the recording are discussed, and methods for compensating for some of these effects are presented. Looking at the distributions of the normalized bit intensities, we are able to estimate the minimum achievable bit error rate for this system.

Three-dimensional optical data storage in a fluorescent dye-doped photopolymer

We propose a new, to our knowledge, monolithic multilayer optical storage medium in which data may be stored through the diffusional redistribution of fluorescent molecules within a polymer host. The active portion of the medium consists of a photopolymer doped with a fluorescent dye that is polymerized at the focal point of a high-numerical-aperture lens. We believe that as fluorescent molecules bond to the polymer matrix they become more highly concentrated in the polymerized regions, resulting in the modulated data pattern. Since data readout is based on detection of fluorescence rather than index modulation as in other photopolymer-based memories, the problems of media shrinkage and optical scatter are of less concern. An intensity threshold observed in the recording response of this material due to the presence of inhibitor molecules in the photopolymer allows for the three-dimensional confinement of recorded bits and therefore multilayer recording. The nonlinear recording characteristics of this material were investigated through a simple model of photopolymerization and diffusion and verified experimentally. Both single-layer and multilayer recordings were demonstrated.

Optical forces for noninvasive cellular analysis

A novel, noninvasive measurement technique for quantitative cellular analysis is presented that utilizes the forces generated by an optical beam to evaluate the physical properties of live cells in suspension. In this analysis, a focused, near-infrared laser line with a high cross-sectional intensity gradient is rapidly scanned across a field of cells, and the interaction of those cells with the beam is monitored. The response of each cell to the laser depends on its size, structure, morphology, composition, and surface membrane properties; therefore, with this technique, cell populations of different type, treatment, or biological state can be compared. To demonstrate the utility of this cell analysis platform, we evaluated the early stages of apoptosis induced in the U937 cancer cell line by the drug camptothecin and compared the results with established reference assays. Measurements on our platform show detection of cellular changes earlier than either of the fluorescence-based Annexin V or caspase assays. Because no labeling or additional cell processing is required and because accurate assays can be performed with a small number of cells, this measurement technique may find suitable applications in cell research, medical diagnostics, and drug discovery.

Three-dimensional holographic stamping of multi-layer bit-oriented non-linear optical media

The requirements and limitations on the use of a volume holographic element for the simultaneous optical stamping of multilayer data into a three-dimensional (3D) bit-oriented material that exhibits a suitable sensitivity threshold are investigated. The expected performance of such a holographic stamping element is examined through a model of the coherent noise effects that result from the interference of the many data layers with one another. We show that higher signal-to-noise values may be achieved through the use of semicoherent light during the readout of the hologram. The main limitations to this technique arise from the bandwidth requirements on the holographic element, the degree of nonlinearity required of the bit-oriented media, and the tolerance requirements for the optical exposure levels. As a demonstration of the concept, a two-layer stamping element is fabricated and used to simultaneously stamp two layers of data into a 3D dye-doped photopolymer storage medium.

Materials and systems for two photon 3-D ROM devices

The methods and systems used for storing and accessing information in three dimensions by means of two-photon absorption are described. The materials into which the information is stored are organic molecules dispersed in polymer matrices, which change structure and spectra after absorption of light. The writing and accessing of the information can be performed either bit-by-bit or in a two-dimensional (2-D) multibit plane format. Automated recording and readout three-dimensional (3-D) systems have been constructed and characterized. Channel error sources have been identified, and a custom spatial bit-error-rate test has been developed.

Cationic two-photon induced polymerization with high dynamic range.

Cationic-induced two-photon photo-polymerization is demonstrated at 710 nm, using an isopropylthioxanthone / diarylidonium salt initiating system for the cationic polymerization of an epoxide. In-situ monitoring of the polymer conversion using interferometry allows for determination of the polymerization threshold J2th, polymerization rate R and its dependence of initiators concentration z. Best J2th achieved is 1 GW/cm 2 , with a dynamic range of > 100, i.e. the material can be fully polymerized at intensities > 100 times the threshold level without damage. The R is found to be proportional to the m=1.7 power of the intensity, or R =[C(J-J2th)]m =[C(J-J2th)]1.7 , which implies a significantly stronger localization of the photochemical response than that of free radical photoinitiators. Both R and J2th significantly improve when the concentration z of the initiator (onium salt) increases, reduction of J2th exhibiting z -m trend.

Integration of optoelectronic array devices for cell transport and sorting

Current biochip technologies typically rely on electrostatic or mechanical forces for the transport and sorting of biological samples such as single cells. In this paper we have investigated how optical pressure forces can be effectively used for the manipulation of cells and switching in a microfluidic system. By projecting the optical beams externally non-contact between the control devices and the sample chip is possible thus allowing the sample chips to be disposable which reduces the chance of cross-contamination. In one implementation we have shown that vertical cavity surface emitting laser (VCSEL) array devices used as parallel optical tweezer arrays can increase the parallelism of sample manipulation on a chip. We have demonstrated the use of a high-order Laguerre-Gaussian mode VCSEL for optical tweezing of polystyrene microspheres and live cells. We have also shown that optical pressure forces from higher- power sources can be used for the switching of particles within microfluidic channels. Both the attractive gradient force and the scattering force of a focused optical beam have been used to direct small particles flowing through junctions molded in PDMS. We believe that by integrating optical array devices for simultaneous detection and manipulation, highly parallel and low-cost analysis and sorting devices may be achieved.