Light-based advanced manufacturing increasingly requires programmable, closed-loop tools that translate human design intent into executable operations at small length scales. Yet a key bottleneck persists across robotic and manufacturing modalities: turning user intent into machine-readable objectives that are reliably executable. While micro-robotics offers versatile manipulation via optical actuation of fluids, mathematically tractable goal specification remains manual and hard to reuse. Here, we introduce Speak-to-Objective, a modular agentic pipeline that uses a conditioned Large Language Model (LLM) to translate spoken or written commands into fully differentiable objective functions for assembling microparticles in a constraint-aware inverse solver (SLSQP) and on an experimental optofluidic platform. The approach employs a compact loop - perceive -> compose -> propose -> act -> report learn - that treats the objective as the interface between intent and actuation, separating what to assemble or pattern from how to actuate, while learning from user feedback. The pipeline composes geometry, spacing, and assignment/topology terms to generate robust descriptive objectives that assemble from partial traces and recover after perturbations, as well as explicit objectives for precise placement, all in an actuator-agnostic fashion. Using laser-induced thermoviscous flows as the physical actuation modality, we demonstrate natural-language-programmable, light-based microscale assembly of particle patterns in a microfluidic environment. Beyond its immediate impact on programmable microassembly, and using laser-induced optofluidic actuation as a reduced-complexity experimental platform, our work points toward self-driving, AI-assisted optical manufacturing platforms in which natural language, differentiable objectives, and laser-based actuation are coupled into a reusable digital workflow.

Determination of the rheological properties of cells is known to require active measurements, which largely depend on the internalization of mechanical probes. Here, we circumvent this problem via the introduction of Rheological focused light-induced cytoplasmic streaming (Rheo-FLUCS): an active, yet probe-free approach that leverages light-induced flows to access mechanical changes in complex systems. While Rheo-FLUCS is facilitated by thermoviscous expansion phenomena rather than external forces, here we show equivalence in its ability to measure relative viscoelastic properties. Specifically, we demonstrate a phase-lag equivalence with probe-dependent active microrheology in a wide range of physically different, yet chemically identical materials. We exemplify the utility of Rheo-FLUCS in three distinctly different biological systems: compound-treated mouse fibroblasts (NIH-3T3), genetically modified human osteoblasts (U2OS) to elucidate the role of myosins in cytoplasmic mechanics, and early ascidian oocytes of Phallusia mammillata at fertilization stage. Our biological use-cases exemplify the application versatility of Rheo-FLUCS, which in the future may use phase information as a marker for developmental success.

Abstract Contact-free and object-agnostic three-dimensional (3D) rotation remains a challenge at both the micro and nanoscale, with broad relevance to advanced imaging, biology, microrobotics, and materials science. Specifically, precise 3D rotation is desirable in diffusion-suppressing environments, where conventional micromanipulation methods fail. Here we introduce an opto-thermoviscous strategy that scans a focused laser spot within a two-dimensional plane to robustly generate 3D helical thermoviscous flows (TVFs) within highly viscous media. We further report on the discovery of opto-hydrodynamic focusing that converges a spiral motion to a defined particle height. By exploiting symmetry relations, we use rational design to decouple out-of-plane rotation from lateral displacements, leading to stable spinning with positional fluctuations below 200 nm, and demonstrate compatibility with a broad range of microstructures, from nano-printed tiles to stained biological cells, and even perfectly round homogenous spheres. Finally, leveraging the kinematic nature of thermoviscous manipulations, we demonstrate how stepwise rotation, alternated with 3D volumetric microscopy, can be combined with established multi-view image fusion strategies to increase resolution in biological imaging. Conceptually, this elevates TVFs from planar transport to symmetry-engineered volumetric actuation, delivering robust, material-agnostic, out-of-plane rotational control and sheathless opto-hydrodynamic focusing for all-optical micromanipulations and augmented microscopy.

High-precision micromanipulation techniques, including optical tweezers and hydrodynamic trapping, have garnered wide-spread interest. Recent advances in optofluidic multiplexed assembly and microrobotics demonstrate significant progress, particularly by iteratively applying laser-induced, localized flow fields to manipulate microparticles in viscous solutions. However, these approaches still face challenges such as undesired hydrodynamic coupling and instabilities when multiple particles are brought into close proximity. By leveraging an analytical model of thermoviscous flows, this work introduces a stochastic optimization approach that selects flow fields for precise particle arrangement without relying on rule-based heuristics. Through minimizing a comprehensive objective function, the method achieves sub-micrometer alignment accuracy even in a crowded setting, avoiding instabilities driven by undesired coupling or particle collisions. An autonomously emerging "action at a distance" strategy-placing the laser scan path farther from the manipulated particles over time-exploits the 1/r2 decay of thermoviscous flow to refine positioning. Overall, objective function-based model predictive control enhances the versatility of automated optofluidic manipulations, opening new avenues in assembly, micromanufacturing, robotics, and life sciences.

The nucleus is highly organized to facilitate coordinated gene transcription. Measuring the rheological properties of the nucleus and its sub-compartments will be crucial to understand the principles underlying nuclear organization. Here, we show that strongly localized temperature gradients (approaching 1°C/µm) can lead to substantial intra-nuclear chromatin displacements (>1 µm), while nuclear area and lamina shape remain unaffected. Using particle image velocimetry (PIV), intra-nuclear displacement fields can be calculated and converted into spatio-temporally resolved maps of various strain components. Using this approach, we show that chromatin displacements are highly reversible, indicating that elastic contributions are dominant in maintaining nuclear organization on the time scale of seconds. In genetically inverted nuclei, centrally compacted heterochromatin displays high resistance to deformation, giving a rigid, solid-like appearance. Correlating spatially resolved strain maps with fluorescent reporters in conventional interphase nuclei reveals that various nuclear compartments possess distinct mechanical identities. Surprisingly, both densely and loosely packed chromatin showed high resistance to deformation, compared to medium dense chromatin. Equally, nucleoli display particularly high resistance and strong local anchoring to heterochromatin. Our results establish how localized temperature gradients can be used to drive nuclear compartments out of mechanical equilibrium to obtain spatial maps of their material responses.

Techniques for high-definition micromanipulations, such as optical tweezers, hold substantial interest across a wide range of disciplines. However, their applicability remains constrained by material properties and laser exposure. And while microfluidic manipulations have been suggested as an alternative, their inherent capabilities are limited and further hindered by practical challenges of implementation and control. Here we show that the iterative application of laser-induced, localized flow fields can be used for the relative positioning of multiple micro-particles, irrespectively of their material properties. Compared to the standing theoretical proposal, our method keeps particles mobile, and we show that their precision manipulation is non-linearly accelerated via the multiplexing of temperature stimuli below the heat diffusion limit. The resulting flow fields are topologically rich and mathematically predictable. They represent unprecedented microfluidic control capabilities that are illustrated by the actuation of humanoid micro-robots with up to 30 degrees of freedom, whose motions are sufficiently well-defined to reliably communicate personal characteristics such as gender, happiness and nervousness. Our results constitute high-definition micro-fluidic manipulations with transformative potential for assembly, micro-manufacturing, the life sciences, robotics and opto-hydraulically actuated micro-factories.

During development, the conserved PAR polarity network is continuously redeployed, requiring that it adapt to changing cellular contexts and environmental cues. In the early C. elegans embryo, polarity shifts from being a cell-autonomous process in the zygote to one that must be coordinated between neighbors as the embryo becomes multicellular. Here, we sought to explore how the PAR network adapts to this shift in the highly tractable C. elegans germline P lineage. We find that although P lineage blastomeres exhibit a distinct pattern of polarity emergence compared with the zygote, the underlying mechanochemical processes that drive po-larity are largely conserved. However, changes in the symmetry-breaking cues of P lineage blastomeres ensure coordination of their polarity axis with neighboring cells. Specifically, we show that furrow-directed cortical flows associated with cytokinesis of the zygote induce symmetry breaking in the germline blastomere P1 by transporting PAR-3 into the nascent cell contact. This pool of PAR-3 then biases downstream PAR polarization pathways to establish the polarity axis of P1 with respect to the position of its anterior sister, AB. Thus, our data suggest that cytokinesis itself induces symmetry breaking through the advection of polarity proteins by furrow-directed flows. By directly linking cell polarity to cell division, furrow-directed cortical flows could be a general mechanism to ensure proper organization of cell polarity within actively dividing systems.

Recently, it has been demonstrated that thermoviscous flows can be used for a range of fine micromanipulations, such as moving the cytoplasm of cells and developing embryos, intracellular rheology, and femtonewton-range force measurements. These flows, also known as focused-light-induced cytoplasmic streaming (FLUCS), are induced by mid-infrared laser scanning of a temperature spot through the sample. However, localized laser scanning can inflict temperature perturbations of several Kelvins on the sample, potentially eliciting unspecific biological responses. In this study, we demonstrate how exploiting symmetry relations during laser scanning effectively disentangles laser heating and flow induction. We introduce flow-neutral scan sequences that use dynamic photothermal stimuli and spatiotemporal symmetry relations of scanning bridging up to three distinct time scales. We leverage further insights from a recently published analytical model of flow fields to present quasi-homogenous temperature distributions that leave flow lines and their local and directed character largely invariant. We present practical, intuitive solutions through predesigned sets of scan lines with near isothermal distributions and demonstrate that they are sufficient to generate and control flows in Caenorhabditis elegans embryos on a magnitude well in excess of endogenous flow velocities. Our results enable the separation of two previously tightly linked classes of physical stimuli, introduce a new, even less invasive standard for performing FLUCS perturbations, and pave the way for new unexplored avenues in the fields of soft matter and biomedicine.

Recent experiments in cell biology have probed the impact of artificially-induced intracellular flows in the spatiotemporal organisation of cells and organisms. In these experiments, mild dynamical heating via focused infrared light from a laser leads to long-range, thermoviscous flows of the cytoplasm inside a cell, a method popularised in cell biology as FLUCS (focused-light-induced cytoplasmic streaming). Here, we present a fully analytical, theoretical model describing the fluid flow and transport of tracers induced by the laser at all length scales in two-dimensional confinement. The focused light causes a small, local temperature change, which in turn results in a small change in the density and viscosity of the fluid locally. We analytically solve for the instantaneous flow field due to the translation of a heat spot of arbitrary time-dependent amplitude along a scan path. We show that the leading-order instantaneous flow results purely from thermal expansion. It is proportional to the magnitude of the temperature perturbation, with far-field behaviour typically dominated by a source or sink flow and proportional to the rate of change of the heat-spot amplitude. In contrast, and in agreement with experimental measurements, the net displacement of a tracer due to a full scan of the heat spot is quadratic in the heat-spot amplitude, as it results from the interplay of thermal expansion and thermal viscosity changes. The corresponding average velocity of tracers over a scan is a hydrodynamic source dipole in the far field, with direction dependent on the relative importance of thermal expansion and thermal viscosity changes. Our quantitative findings show excellent agreement with recent experimental results and will enable the design of new controlled experiments to establish the physiological role of physical transport processes inside cells.

It has long been proposed that phase-separated compartments can provide a basis for the formation of cellular precursors in prebiotic environments. However, we know very little about the properties of coacervates formed from simple peptides, their compatibility with ribozymes or their functional significance. Here we assess the conditions under which functional ribozymes form coacervates with simple peptides. We find coacervation to be most robust when transitioning from long homopeptides to shorter, more pre-biologically plausible heteropeptides. We mechanistically show that these RNA-peptide coacervates display peptide-dependent material properties and cofactor concentrations. We find that the interspacing of cationic and neutral amino acids increases RNA mobility, and we use isothermal calorimetry to reveal sequence-dependent Mg2+ partitioning, two critical factors that together enable ribozyme activity. Our results establish how peptides of limited length, homogeneity and charge density facilitate the compartmentalization of active ribozymes into non-gelating, magnesium-rich coacervates, a scenario that could be applicable to cellular precursors with peptide-dependent functional phenotypes.

Microscopic flows and the associated transport of biological components inside cells play an important role in the development and homeostasis of living cells. A longstanding question is how molecules suddenly self-organise into spatial structures like e.g. biological cells. Until recently, it was not possible to generate such active flows inside biological cells in a controlled manner. With FLUCS—a method developed by the Kreysing lab—it is possible to investigate physical principles of cell organization.

Membraneless compartments, also known as condensates, provide chemically distinct environments and thus spatially organize the cell. A well-studied example of condensates is P granules in the roundworm Caenorhabditis elegans that play an important role in the development of the germline. P granules are RNA-rich protein condensates that share the key properties of liquid droplets such as a spherical shape, the ability to fuse, and fast diffusion of their molecular components. An outstanding question is to what extent phase separation at thermodynamic equilibrium is appropriate to describe the formation of condensates in an active cellular environment. To address this question, we investigate the response of P granule condensates in living cells to temperature changes. We observe that P granules dissolve upon increasing the temperature and recondense upon lowering the temperature in a reversible manner. Strikingly, this temperature response can be captured by in vivo phase diagrams that are well described by a Flory-Huggins model at thermodynamic equilibrium. This finding is surprising due to active processes in a living cell. To address the impact of such active processes on intracellular phase separation, we discuss temperature heterogeneities. We show that, for typical estimates of the density of active processes, temperature represents a well-defined variable and that mesoscopic volume elements are at local thermodynamic equilibrium. Our findings provide strong evidence that P granule assembly and disassembly are governed by phase separation based on local thermal equilibria where the nonequilibrium nature of the cytoplasm is manifested on larger scales.

Microscale Janus emulsions represent a versatile material platform for dynamic refractive, reflective, and light-emitting optical components. Here, we present a mechanism for droplet actuation that exploits thermocapillarity. Using optically induced thermal gradients, an interfacial tension differential is generated across the surfactant-free internal capillary interface of Janus droplets. The interfacial tension differential causes droplet-internal Marangoni flows and a net torque, resulting in a predictable and controllable reorientation of the droplets. The effect can be quantitatively described with a simple model that balances gravitational and thermal torques. Occurring in small thermal gradients, these optothermally induced Marangoni dynamics represent a promising mechanism for controlling droplet-based micro-optical components.

Oocytes are large cells that develop into an embryo upon fertilization 1 . As interconnected germ cells mature into oocytes, some of them grow—typically at the expense of others that undergo cell death 2 – 4 . We present evidence that in the nematode Caenorhabditis elegans , this cell-fate decision is mechanical and related to tissue hydraulics. An analysis of germ cell volumes and material fluxes identifies a hydraulic instability that amplifies volume differences and causes some germ cells to grow and others to shrink, a phenomenon that is related to the two-balloon instability 5 . Shrinking germ cells are extruded and they die, as we demonstrate by artificially reducing germ cell volumes via thermoviscous pumping 6 . Our work reveals a hydraulic symmetry-breaking transition central to the decision between life and death in the nematode germline.

The ability to control the position of micron-size particles with high precision using tools such as optical tweezers has led to major advances in fields such as biology, physics and material science. In this paper, we present a novel optical strategy to confine particles in solution with high spatial control using feedback-controlled thermoviscous flows. We show that this technique allows micron-size particles to be positioned and confined with subdiffraction precision (24 nm), effectively suppressing their diffusion. Due to its physical characteristics, our approach might be particular attractive where laser exposure is of concern or materials are inherently incompatible with optical tweezing since it does not rely on contrast in the refractive index.

The use of optical tweezers to measure forces acting upon microscopic particles has revolutionised fields from material science to cell biology. However, despite optical control capabilities, this technology is highly constrained by the material properties of the probe, and its use may be limited due to concerns about the effect on biological processes. Here we present a novel, optically controlled trapping method based on light-induced hydrodynamic flows. Specifically, we leverage optical control capabilities to convert a translationally invariant topological defect of a flow field into an attractor for colloids in an effectively one-dimensional harmonic, yet freely rotatable system. Circumventing the need to stabilise particle dynamics along an unstable axis, this novel trap closely resembles the isotropic dynamics of optical tweezers. Using magnetic beads, we explicitly show the existence of a linear force-extension relationship that can be used to detect femtoNewton-range forces with sensitivity close to the thermal limit. Our force measurements remove the need for laser-particle contact, while also lifting material constraints, which renders them a particularly interesting tool for the life sciences and engineering.

Mutations in cis-regulatory elements play important roles for phenotypic changes during evolution. Eye degeneration in the blind mole rat (BMR; Nannospalax galili) and other subterranean mammals is significantly associated with widespread divergence of eye regulatory elements, but the effect of these regulatory mutations on eye development and function has not been explored. Here, we investigate the effect of mutations observed in the BMR sequence of a conserved noncoding element upstream of Tdrd7, a pleiotropic gene required for lens development and spermatogenesis. We first show that this conserved element is a transcriptional repressor in lens cells and that the BMR sequence partially lost repressor activity. Next, we recapitulated evolutionary changes in this element by precisely replacing the endogenous regulatory element in a mouse line by the orthologous BMR sequence with CRISPR-Cas9. Strikingly, this repressor replacement caused a more than 2-fold upregulation of Tdrd7 in the developing lens; however, increased mRNA level does not result in a corresponding increase in TDRD7 protein nor an obvious lens phenotype, possibly explained by buffering at the posttranscriptional level. Our results are consistent with eye degeneration in subterranean mammals having a polygenic basis where many small-effect mutations in different eye-regulatory elements collectively contribute to phenotypic differences.

ABSTRACTThe nucleus is highly organized to facilitate coordinated gene transcription. Measuring the rheological properties of the nucleus and its sub-compartments will be crucial to understand the principles underlying nuclear organization. Here, we show that strongly localized temperature gradients (approaching 1°C /μm) can lead to substantial intra-nuclear chromatin displacements (>1 μm), while nuclear area and lamina shape remain unaffected. Using particle image velocimetry (PIV), intra-nuclear displacement fields can be calculated and converted into spatio-temporally resolved maps of various strain components. Using this approach, we show that chromatin displacements are highly reversible, indicating that elastic contributions are dominant in maintaining nuclear organization on the time scale of seconds. In genetically inverted nuclei, centrally compacted heterochromatin displays high resistance to deformation, giving a rigid, solid-like appearance. Correlating spatially resolved strain maps with fluorescent reporters in conventional interphase nuclei reveals that various nuclear compartments possess distinct mechanical identities. Surprisingly, both densely and loosely packed chromatin showed high resistance to deformation, compared to medium dense chromatin. Equally, nucleoli display particularly high rigidity and strong local anchoring to heterochromatin. Our results establish how localized temperature gradients can be used to drive nuclear compartments out of mechanical equilibrium to obtain spatial maps of their material responses.Main FindingsNovel non-invasive active micro-rheology method to probe spatial intranuclear material responses, unhindered by the nuclear lamina, using strongly localized temperature gradientsChromatin shows both elastic and viscous properties at the mesoscale with a retardation time of τ ∼ 1sCompacted heterochromatin in a model of nuclear inversion shows high resistance to deformation, suggesting dominantly solid-like behaviorThe nucleus displays spatially distinct material properties for different compartmentsThe nucleolus shows high resistance to deformation on the time scale of secondsImmobile nucleoli appear solidly anchored to and retain the deformation of surrounding chromatin

Membraneless compartments, also known as condensates, provide chemically distinct environments and thus spatially organize the cell. A well-studied example of condensates is P granules in the roundworm C. elegans which play an important role in the development of the germline. P granules are RNA-rich protein condensates that share the key properties of liquid droplets such as a spherical shape, the ability to fuse, and fast diffusion of their molecular components. An outstanding question is to what extent phase separation at thermodynamic equilibrium is appropriate to describe the formation of condensates in an active cellular environment. To address this question, we investigate the response of P granule condensates in living cells to temperature changes. We observe that P granules dissolve upon increasing the temperature and recondense upon lowering the temperature in a reversible manner. Strikingly, this temperature response can be captured by in vivo phase diagrams which are well described by a Flory-Huggins model at thermodynamic equilibrium. This finding is surprising due to active processes in a living cell. To address the impact of such active processes on intra-cellular phase separation, we discuss temperature heterogeneities. We show that, for typical estimates of the density of active processes, temperature represents a well-defined variable and that mesoscopic volume elements are at local thermodynamic equilibrium. Our findings provide strong evidence that P granule assembly and disassembly are governed by phase separation based on local thermal equilibria where the non-equilibrium nature of the cytoplasm is manifested on larger scales. SIGNIFICANCE STATEMENT Living cells rely on a continuous flux of energy to spatially organize biochemical processes. It remained unclear whether cells can achieve this spatial organization via thermodynamic principles. Here, we report the striking behavior of a cold-blooded organism that reacts to environmental temperature changes similar to a thermodynamic system at local equilibrium. Our key finding is that protein-rich droplets form and dissolve reversibly with temperature due to changes in the organism?s entropy. We show that the organism uses a specific molecule to extend droplet stability to the natural temperature range of the organism’s habitat. Due to the relevance of such protein droplets for the organism?s fertility, our works shed light on how molecular components could facilitate biological functions via thermodynamic principles.