# Non-invasive measurement of biomolecular condensate interfacial tension and bending rigidity

## TL;DR

A new open-source tool called FlickerPrint measures mechanical properties of biomolecular condensates and vesicles using shape fluctuations in microscopy images.

## Contribution

FlickerPrint enables high-throughput, non-invasive measurement of interfacial tension and bending rigidity of soft biological structures.

## Key findings

- FlickerPrint can analyze thousands of condensates and vesicles using confocal microscopy data.
- The method is robust to changes in imaging setup, including frame rate.
- The tool is applicable to live cells and in vitro systems.

## Abstract

Accurate measurement of biomolecular condensates’ mechanical properties is essential to understand their behavior within cells. We present FlickerPrint, an open-source Python package to determine the interfacial tension and bending rigidity of thousands of condensates using flicker spectroscopy by analyzing their shape fluctuations in confocal microscopy images. We detail the workflow and computational requirements of FlickerPrint to scale up these individual measurements to the population level. Examples of experiments in live cells and in vitro that are suitable for analysis with FlickerPrint are provided, as well as scenarios where the package cannot be used. Using these examples, we show that the results obtained are robust to changes in imaging setup, including frame rate. This implementation enables a step change in measurement capability for two key properties of biomolecular condensates: interfacial tension and bending rigidity. Moreover, the tools in FlickerPrint are also applicable for analyzing other soft, fluctuating bodies, demonstrated here using vesicles.

•We present a Python package to infer mechanical properties of condensates and vesicles•The method is non-invasive and is applicable to bodies in live cells and in vitro•Interfacial tension and bending rigidity are measured using high-throughput analysis•We provide examples of suitable systems and considerations for imaging

We present a Python package to infer mechanical properties of condensates and vesicles

The method is non-invasive and is applicable to bodies in live cells and in vitro

Interfacial tension and bending rigidity are measured using high-throughput analysis

We provide examples of suitable systems and considerations for imaging

Biomolecular condensates play fundamental roles in sub-cellular organization, and it is well known that the composition of condensates can affect their function. Measuring the condensates’ mechanical properties (for example, interfacial tension and bending rigidity) can aid the understanding of their biomolecular composition and cellular functions. However, measuring the properties of individual condensates under physiological conditions is very challenging and cumbersome to scale to the population level using traditional methods. To overcome these issues, we have developed a software package to run flicker spectroscopy analysis of condensates at scale, to determine their interfacial tension and bending rigidity. At the same time, the package can be harnessed to analyze other soft, fluctuating bodies such as lipid vesicles.

Williamson et al. present FlickerPrint, an open-source computational analysis tool for measuring the interfacial tension and bending rigidity of soft bodies from their shape fluctuations in time-series confocal microscopy videos. The package allows for thousands of biomolecular condensates, vesicles, and other soft bodies to be analyzed.