Measuring condensate material properties

Biomolecular condensates are membraneless organelles central to cellular organization, but connecting their biological function to their material properties such as viscosity, surface tension, and viscoelasticity requires quantitative measurements in realistic geometries. Most existing methods identify only one material property, or at best ratios between two properties.

In (Roggeveen et al., 2023), we developed a calibration-free model of micropipette aspiration for liquid-like protein condensates. Micropipette aspiration can in principle measure both viscosity and surface tension, but models inherited from membrane-bound objects do not correctly describe unbounded liquid condensates. We derived a hydrodynamic model matched to the aspiration geometry, validated it on silicone oil of known properties, and used it to reinterpret measurements of LAF-1 RGG condensates. Our results indicated LAF-1 RGG has a viscosity of 11±1.1 Pa·s and a surface tension of 0.17±0.02 mN/m.

Micropipette aspiration provides a direct mechanical perturbation of small liquid-like condensates. Experiments courtesy of Huan Wang.

By plotting pressure drop against the model fitting parameter, we extract viscosity from the slope and surface tension from the scaled reciprocal intercept.

Collapse of silicone-oil aspiration curves under the calibration-free hydrodynamic model using our experimentally derived material parameters.

Collapsed aspiration curves for LAF-1 RGG condensates using the same model framework.

Aspiration of two-phase nucleoli in a near-native cellular environment. Purple marks the granular component; green marks the fibrillar component. Experiment by Holly Cheng.

The second part of this line of work applied micropipette aspiration to nucleoli in a near-native cellular environment (Cheng et al., 2025). The measurements showed that nucleolar subcompartments have distinct material properties: the outer granular component behaves more fluid-like, while the inner dense fibrillar component is more viscoelastic and depends strongly on RNA.

Because of the wetting properties of these condensates, we could not develop a complete hydrodynamic model. However, we could still show that the elastic response of the fibrillar component arises from cross-linking by long RNA molecules. A viscoelastic solid responds qualitatively differently to aspiration than a fluid; after RNA cleavage, the dense fibrillar component shifted toward a more fluid-like response.

Why this matters

Condensates are often described qualitatively as liquid-like or solid-like, but biological function depends on quantitative material properties. This work connects experimental aspiration data to interpretable mechanical models, making it possible to compare viscosity, surface tension, and viscoelastic response across systems and connect those properties to biological function.

Schematic of the response of different idealized, one-dimensional materials to a stress profile consistent with micropipette aspiration. The fluid-like models (Newtonian, Maxwell) show a linear increase in strain over time, while the solid model (Kelvin-Voigt) saturates at a fixed value.

Untreated nucleoli show distinct mechanical responses across subcompartments. The purple granular component resembles that of a Newtonian fluid while the response of the green fibrillar component resembles a viscoelastic solid.

Treated nucleoli show altered aspiration dynamics after perturbing RNA-dependent material structure. Now both materials respond like a fluid.

References

2025

Micropipette aspiration reveals differential RNA-dependent viscoelasticity of nucleolar subcompartments

Holly H. Cheng , James V. Roggeveen , Huan Wang , and 3 more authors

Proceedings of the National Academy of Sciences, May 2025

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The nucleolus is a multiphasic biomolecular condensate that facilitates ribosome biogenesis, a complex process involving hundreds of proteins and RNAs. The proper execution of ribosome biogenesis likely depends on the material properties of the nucleolus. However, these material properties remain poorly understood due to the challenges of in vivo measurements. Here, we use micropipette aspiration (MPA) to directly characterize the viscoelasticity and interfacial tensions of nucleoli within transcriptionally active Xenopus laevis oocytes. We examine the major nucleolar subphases, the outer granular component (GC) and the inner dense fibrillar component (DFC), which itself contains a third small phase known as the fibrillar center (FC). We show that the behavior of the GC is more liquid-like, while the behavior of the DFC/FC is consistent with that of a partially viscoelastic solid. To determine the role of ribosomal RNA in nucleolar material properties, we degrade RNA using RNase A, which causes the DFC/FC to become more fluid-like and alters interfacial tension. Together, our findings suggest that RNA underlies the partially solid-like properties of the DFC/FC and provide insights into how material properties of nucleoli in a near-native environment are related to their RNA-dependent function.

2023

A calibration-free model of micropipette aspiration for measuring properties of protein condensates

James V. Roggeveen , Huan Wang , Zheng Shi , and 1 more author

Biophysical Journal, Oct 2023

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There is growing evidence that biological condensates, which are also referred to as membraneless organelles, and liquid-liquid phase separation play critical roles regulating many important cellular processes. Understanding the roles these condensates play in biology is predicated on understanding the material properties of these complex substances. Recently, micropipette aspiration (MPA) has been proposed as a tool to assay the viscosity and surface tension of condensates. This tool allows the measurement of both material properties in one relatively simple experiment, in contrast to many other techniques that only provide one or a ratio of parameters. While this technique has been commonly used in the literature to determine the material properties of membrane-bound objects dating back decades, the model describing the dynamics of MPA for objects with an external membrane does not correctly capture the hydrodynamics of unbounded fluids, leading to a calibration parameter several orders of magnitude larger than predicted. In this work we derive a new model for MPA of biological condensates that does not require any calibration and is consistent with the hydrodynamics of the MPA geometry. We validate the predictions of this model by conducting MPA experiments on a standard silicone oil of known material properties and are able to predict the viscosity and surface tension using MPA. Finally, we reanalyze with this new model the MPA data presented in previous works for condensates formed from LAF-1 RGG domains.