Current optically actuated microrobots have functionalities that are mostly mechanical in nature. These microrobots serve to mediate and relay optical forces, which is particularly useful for objects that cannot be trapped directly due to poor index contrast, incompatible size and shape or to prevent unwanted reactions with light. Optical forces are weak in general but light can be used in a clever way to initiate other effects to supplement optical trapping. The next generation of microrobots covered in this chapter demonstrate new functionalities, such as enhanced photothermal heating to generate convection currents to draw in cargo, enhanced signals for spectroscopy, and localized temperature sensing. The new microrobots require additional fabrication steps to integrate metallic structures to the usual polymer body. Metals are very good at converting optical energy to heat, can have high thermal conductivities and can enhance nonlinear effects. Integrating all this in a light-controlled vehicle can bring about the aforementioned functionalities with precise spatial and temporal control. We envision these next generation light-driven microrobots to be an important addition to our Light Robotics toolbox in understanding biology at the micro- and nanoscales.

Recently we proposed the concept of so-called Light Robotics including the new and disruptive 3Dfabricated micro-tools coined Wave-guided Optical Waveguides that can be real-time optically manipulated and remote-controlled with a joystick in a volume with six-degrees-of-freedom. Exploring the full potential of this new ` drone-like' light-driven micro-robotics in challenging microscopic geometries requires a versatile and real-time reconfigurable light addressing that can dynamically track a plurality of tiny micro-robots in 3D to ensure continuous optimal light coupling on the fly. Our latest developments in this new and exciting research area will be reviewed.

Light can be used to fabricate, handle, power, and actuate microrobotics functionalities, such as the loading and unloading of micro-cargo, showing promise for drug delivery and biological-testing applications.

Early detection of diseases can save lives. Hence, there is emphasis in sorting rare disease-indicating cells within small dilute quantities such as in the confines of lab-on-a-chip devices. However, before diseased cells can be studied in isolation, it is necessary to identify them against normal healthy cells. With the richness of visual information, a lot of microscopy techniques have been developed and have been crucial in biological studies. To utilize their complementary advantages we adopt both fluorescence and brightfield imaging in our optical cell sorter. Brightfield imaging has the advantage of being non-invasive, thus maintaining cell viability. Fluorescence imaging, on the other hand, takes advantages of the chemical specificity of fluorescence markers and can validate machine vision results from brightfield images. Visually identified cells are sorted using optical manipulation techniques. Scattering forces from beams actuated via efficient phase-only efficient modulation has been adopted. This has lowered the required power for sorting cells to a tenth of our previous approach, and also makes the cell sorter safer for use in clinical settings. With the versatility of dynamically programmable phase spatial light modulators, a plurality of light shaping techniques, including hybrid approaches, can be utilized in cell sorting.

Light Robotics is a new field of research where ingredients from photonics, nanotechnology and biotechnology are put together in new ways to realize light-driven robotics at the smallest scales to solve major challenges primarily within the nanobio-domain but not limited hereto. Exploring the full potential of this new 'drone-like' light-printed, light-driven, light-actuated micro-and nanorobotics in challenging geometries requires a versatile and real-time reconfigurable light addressing that can dynamically track a plurality of tiny tools in 3D to ensure real-time continuous light-delivery on the fly. Our latest developments in this new and exciting research area will be reviewed.

Material transport is an important mechanism in microfluidics and drug delivery. The methods and solutions found in literature involve passively diffusing structures, microneedles and chemically fueled structures. In this work, we make use of optically actuated microtools with embedded metal layer as heating element for controlled loading and release. The new microtools take advantage of the photothermal-induced convection current to load and unload cargo. We also discuss some challenges encountered in realizing a self-contained polymerized microtool. Microfluidic mixing, fluid flow control and convection currents have been demonstrated both experimentally and numerically for static metal thin films or passively floating nanoparticles. Here we show an integration of aforementioned functionalities in an optically fabricated and actuated microtool. As proof of concept, we demonstrate loading and unloading of beads. This can be extended to controlled transport and release of genetic material, bio-molecules, fluorescent dyes. We envisioned these microtools to be an important addition to the portfolio of structure-mediated contemporary biophotonics.

Leveraging developments in microfabrication open new possibilities for optical manipulation. With the structural design freedom from three-dimensional printing capabilities of two-photon polymerization, we are starting to see the emergence of cleverly shaped 'light robots' or optically actuated micro-tools that closely resemble their macroscopic counterparts in function and sometimes even in form. In this work, we have fabricated a new type of light robot that is capable of loading and unloading cargo using photothermally induced convection currents within the body of the tool. We have demonstrated this using silica and polystyrene beads as cargo. The flow speeds of the cargo during loading and unloading are significantly larger than when using optical forces alone. This new type of light robot presents a mode of material transport that may have a significant impact on targeted drug delivery and nanofluidics injection.

As celebrated by the Nobel Prize 2014 in Chemistry light-based technologies can now overcome the diffraction barrier for imaging with nanoscopic resolution by so-called super-resolution microscopy1. However, interactive investigations coupled with advanced imaging modalities at these small scale domains gradually demand the development of a new generation of disruptive tools, not only for passively observing at nanoscopic scales, but also for actively reaching into and effectively handling constituents in this size domain. This intriguing mindset has recently led to the emergence of a novel research discipline that could potentially be able to offer the full packet needed for true "active nanoscopy" by use of so-called light-driven micro-robotics or Light Robotics in short.

Generalized Phase Contrast (GPC) is an efficient method for efficiently shaping light into speckle-free contiguous optical distributions useful in diverse applications such as static beam shaping, optical manipulation and recently, for excitation in two-photon optogenetics. GPC typically results in a 3x intensified user defined input mask shape against a dark background. In this work, we emphasize GPC’s capability of optimal destructive interference, normally used to create the dark background surrounding the shaped light. We also study input parameters wherein the locations of light and darkness are interchanged with respect to typical GPC output, thus resulting to a well-defined structured darkness. The conditions that give destructive interference for the output are then applied to near-arbitrary shapes. Preliminary experimental results are presented using dynamic spatial light modulator to form scaled arbitrary darkness shapes. Supporting demonstrations that reverse the light and dark regions of amplitude-modulated input are also presented as a related case of structuring destructive interference. Our analysis and experimental demonstrations show a simplified approach in the generation of extended regions of destructive interference within coherent beams.

Generalized Phase Contrast (GPC) is a light efficient method for generating speckle-free contiguous optical distributions using binary-only or analog phase levels. It has been used in applications such as optical trapping and manipulation, active microscopy, structured illumination, optical security, parallel laser marking and labelling and recently in contemporary biophotonics applications such as for adaptive and parallel two-photon optogenetics and neurophotonics. We will present our most recent GPC developments geared towards these applications. We first show a very compact static light shaper followed by the potential of GPC for biomedical and multispectral applications where we experimentally demonstrate the active light shaping of a supercontinuum laser over most of the visible wavelength range. Finally, we discuss how GPC can be advantageously applied for Quantitative Phase Imaging (QPI).

We show a simplified method of generating extended regions of destructive interference with near arbitrary shapes using the generalized phase contrast (GPC) method. For Gaussian input beams, GPC typically results in a 3x intensified user-defined input mask shape against a dark background. In this work, we investigate conditions wherein GPC's synthetic reference wave destructively interferes with what is typically the foreground pattern. Using alternate conditions for the input phase mask, the locations of light and darkness are interchanged with respect to typical GPC output mappings. We show experimentally how "dark GPC" allows the dark regions to be easily reshaped using a binary-only phase mask encoded on a spatial light modulator. Similar to standard GPC, the method does not require complex calculations or the fabrication of complex gray-level phase elements. The simplified approach and flexibility in the output shapes make dark GPC attractive for applications such as optical trapping of low-index particles or superresolution microscopy like stimulated emission depletion. (C) The Authors. Published by SPIE

Early detection of diseases can save lives. Hence, there is emphasis in sorting rare disease-indicating cells within small dilute quantities such as in the confines of lab-on-a-chip devices. In our work, we use optical forces to isolate red blood cells detected by machine vision. This approach is gentler, less invasive and more economical compared to conventional FACS systems. As cells are less responsive to plastic or glass beads commonly used in the optical manipulation literature, and since laser safety would be an issue in clinical use, we develop efficient approaches in utilizing lasers and light modulation devices. The Generalized Phase Contrast (GPC) method that can be used for efficiently illuminating spatial light modulators or creating well-defined contiguous optical traps is supplemented by diffractive techniques capable of integrating the available light and creating 2D or 3D beam distributions aimed at the positions of the detected cells. Furthermore, the beam shaping freedom provided by GPC can allow optimizations in the beam’s propagation and its interaction with the catapulted cells.

We have previously demonstrated on-demand dynamic coupling to optically manipulated microtools coined as wave-guided optical waveguides using diffractive techniques on a "point and shoot" approach. These microtools are extended microstructures fabricated using two-photon photopolymerization and function as free-floating optically trapped waveguides. Dynamic coupling of focused light via these structures being moved in three-dimensional space is done holographically. However, calculating the necessary holograms is not straightforward when using counter-propagating trapping geometry. The generation of the coupling spots is done in real time following the position of each microtool with the aid of an object tracking routine. This approach allows continuous coupling of light through the microtools which can be useful in a variety of biophotonics applications. To complement the targeted-light delivery capability of the microtools, the applied spatial light modulator has been illuminated with a properly matched input beam cross section based on the generalized phase contrast method. Our results show a significant gain in the output at the tip of each microtool as measured from the fluorescence signal of the trapping medium. The ability to switch from on-demand to continuous addressing with efficient illumination leverages our microtools for potential applications in stimulation and near-field-based biophotonics on cellular scales. (C) The Authors. Published by SPIE under a Creative Commons Attribution 3.0 Unported License.