Biocompatible micro tweezers for 3D hydrogel organoid array mechanical characterization.

This study presents novel biocompatible Polydimethylsiloxane (PDMS)-based micromechanical tweezers (µTweezers) capable of the stiffness characterization and manipulation of hydrogel-based organoids. The system showed great potential for complementing established mechanical characterization methods such as Atomic Force Microscopy (AFM), parallel plate compression (PPC), and nanoindentation, while significantly reducing the volume of valuable hydrogels used for testing. We achieved a volume reduction of ~0.22 µl/sample using the µTweezers vs. ~157 µl/sample using the PPC, while targeting high-throughput measurement of widely adopted micro-mesoscale (a few hundred µm-1500 µm) 3D cell cultures. The µTweezers applied and measured nano-millinewton forces through cantilever' deflection with high linearity and tunability for different applications; the assembly is compatible with typical inverted optical microscopes and fit on standard tissue culture Petri dishes, allowing mechanical compression characterization of arrayed 3D hydrogel-based organoids in a high throughput manner. The average achievable output per group was 40 tests per hour, where 20 organoids and 20 reference images in one 35 mm petri dish were tested, illustrating efficient productivity to match the increasing demand on 3D organoids' applications. The changes in stiffness of collagen I hydrogel organoids in four conditions were measured, with ovarian cancer cells (SKOV3) or without (control). The Young's modulus of the control group (Control-day 0, E = 407± 146, n = 4) measured by PPC was used as a reference modulus, where the relative elastic compressive modulus of the other groups based on the stiffness measurements was also calculated (control-day 0, E = 407 Pa), (SKOV3-day 0, E = 318 Pa), (control-day 5, E = 528 Pa), and (SKOV3-day 5, E = 376 Pa). The SKOV3-embedded hydrogel-based organoids had more shrinkage and lowered moduli on day 0 and day 5 than controls, consistently, while SKOV3 embedded organoids increased in stiffness in a similar trend to the collagen I control from day 0 to day 5. The proposed method can contribute to the biomedical, biochemical, and regenerative engineering fields, where bulk mechanical characterization is of interest. The µTweezers will also provide attractive design and application concepts to soft membrane-micro 3D robotics, sensors, and actuators.

Brightfield, fluorescence, and phase-contrast whole slide imaging via dual-LED autofocusing.

Whole slide imaging (WSI) systems convert the conventional biological samples into digital images. Existing commercial WSI systems usually require an expensive high-performance motorized stage to implement the precise mechanical control, and the cost is prohibitive for most individual pathologists. In this work, we report a low-cost WSI system using the off-the-shelf components, including a computer numerical control (CNC) router, a photographic lens, a programmable LED array, a fluorescence filter cube, and a surface-mount LED. To perform real-time single-frame autofocusing, we exploited two elements of a programmable LED array to illuminate the sample from two different incident angles. The captured image would contain two copies of the sample with a certain separation determined by the defocus distance of the sample. Then the defocus distance can be recovered by identifying the translational shift of the two copies. The reported WSI system can reach a resolution of ∼0.7 µm. The time to determine the optimal focusing position for each tile is only 0.02 s, which is about an 83% improvement compared to our previous work. We quantified the focusing performance on 1890 different tissue tiles. The mean focusing error is ∼0.34 µm, which is well below the ± 0.7 µm depth of field range of our WSI system. The reported WSI system can handle both the semitransparent and the transparent sample, enabling us to demonstrate the implementation of brightfield, fluorescence, and phase-contrast WSI. An automatic digital distortion correction strategy is also developed to avoid the stitching errors. The reported prototype has an affordable cost and can make it broadly available and utilizable for individual pathologists as well as can promote the development of digital pathology.

Design, Fabrication, and Validation of a Petri Dish-Compatible PDMS Bioreactor for the Tensile Stimulation and Characterization of Microtissues.

In this paper, we report on a novel biocompatible micromechanical bioreactor (actuator and sensor) designed for the in situ manipulation and characterization of live microtissues. The purpose of this study was to develop and validate an application-targeted sterile bioreactor that is accessible, inexpensive, adjustable, and easily fabricated. Our method relies on a simple polydimethylsiloxane (PDMS) molding technique for fabrication and is compatible with commonly-used laboratory equipment and materials. Our unique design includes a flexible thin membrane that allows for the transfer of an external actuation into the PDMS beam-based actuator and sensor placed inside a conventional 35 mm cell culture Petri dish. Through computational analysis followed by experimental testing, we demonstrated its functionality, accuracy, sensitivity, and tunable operating range. Through time-course testing, the actuator delivered strains of over 20% to biodegradable electrospun poly (D, L-lactide-co-glycolide) (PLGA) 85:15 non-aligned nanofibers (~91 µm thick). At the same time, the sensor was able to characterize time-course changes in Young's modulus (down to 10-150 kPa), induced by an application of isopropyl alcohol (IPA). Furthermore, the actuator delivered strains of up to 4% to PDMS monolayers (~30 µm thick), simultaneously characterizing their elastic modulus up to ~2.2 MPa. The platform repeatedly applied dynamic (0.23 Hz) tensile stimuli to live Human Dermal Fibroblast (HDF) cells for 12 hours (h) and recorded the cellular reorientation towards two angle regimes, with averages of -58.85° and +56.02°. The device biocompatibility with live cells was demonstrated for one week, with no signs of cytotoxicity. We can conclude that our PDMS bioreactor is advantageous for low-cost tissue/cell culture micromanipulation studies involving mechanical actuation and characterization. Our device eliminates the need for an expensive experimental setup for cell micromanipulation, increasing the ease of live-cell manipulation studies by providing an affordable way of conducting high-throughput experiments without the need to open the Petri dish, reducing manual handling, cross-contamination, supplies, and costs. The device design, material, and methods allow the user to define the operational range based on their targeted samples/application.

Angular light modulator using optical blinds.

Spatial light modulator (SLM) is widely used in imaging applications for modulating light intensity and phase delay. In this paper, we report a novel device concept termed angular light modulator (ALM). Different from the SLM, the reported ALM employs a tunable blind structure to modulate the angular components of the incoming light waves. For spatial-domain light modulation, the ALM can be directly placed in front of an image sensor for selecting different angular light components. In this case, we can sweep the slat angle of the blind structure and capture multiple images corresponding to different perspectives. These images can then be back-projected for 3D tomographic refocusing. By using a fixed slat angle, we can also convert the incident-angle information into intensity variations for wavefront sensing or introduce a translational shift to the defocused object for high-speed autofocusing. For Fourier-domain light modulation, the ALM can be placed at the pupil plane of an optical system for reinforcing the light propagating trajectories. We show that a pupil-plane-modulated system is able to achieve a better resolution for out-of-focus objects while maintaining the same resolution for in-focus objects. The reported ALM can be fabricated on the chip level and controlled by an external magnetic field. It may provide new insights for developing novel imaging and vision devices.