Biology subjects

Lavania, A. A.

Publications and source records attributed to Lavania, A. A..

3 recordsLinked to original sources

bioRxiv2025

Sizing single trapped nanoparticles with interferometric scattering fluctuations

The sizes of nanoparticles significantly influence their properties but are challenging to quantify at the single-particle level. To enable label-free single-nanoparticle detection, interferometric scattering microscopy has emerged as a sensitive modality that exhibits high-precision axial information encoded in the interferometric scattering signal. Here, we demonstrate that single nanoparticles in an interferometric scattering anti-Brownian electrokinetic (ISABEL) trap can be hydrodynamically sized by interferometic scattering contrast fluctuations. The fluctuation timescale is characterized by time autocorrelation analysis and interpreted via a Brownian Dynamics model of the ISABEL trap to provide an estimate of the hydrodynamic diameter of each trapped particle. We verify this behavior for gold and polystyrene bead standards, and demonstrate performance on single carboxysomes, nanoscale carbon fixation compartments from autotrophic bacteria. The exquisite sensitivity of interferometric scattering amplitude fluctuations further equips researchers with a label-free optical platform for increasingly sophisticated nanoparticle analysis. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=99 SRC="FIGDIR/small/696206v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@1632562org.highwire.dtl.DTLVardef@2b625dorg.highwire.dtl.DTLVardef@46d61dorg.highwire.dtl.DTLVardef@933d59_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2022

Exploring masses and internal mass distributions of single carboxysomes in free solution using fluorescence and interferometric scattering in an anti-Brownian trap

Carboxysomes are self-assembled bacterial microcompartments that facilitate carbon assimilation by co-localizing the enzymes of CO2 fixation within a protein shell. These microcompartments can be highly heterogeneous in their composition and filling, so measuring the mass and loading of an individual carboxysome would allow for better characterization of its assembly and function. To enable detailed and extended characterizations of single nanoparticles in solution, we recently demonstrated an improved Interferometric Scattering Anti-Brownian ELectrokinetic (ISABEL) trap, which tracks the position of a single nanoparticle via its scattering of a near-infrared beam and applies feedback to counteract its Brownian motion. Importantly, the scattering signal can be related to the mass of nanoscale proteinaceous objects, whose refractive indices are well-characterized. We calibrate single-particle scattering cross-section measurements in the ISABEL trap and determine individual carboxysome masses in the 50-400 MDa range by analyzing their scattering cross-sections with a core-shell model. We further investigate carboxysome loading by combining mass measurements with simultaneous fluorescence reporting from labeled internal components. This method may be extended to other biological objects, such as viruses or extracellular vesicles, and can be combined with orthogonal fluorescence reporters to achieve precise physical and chemical characterization of individual nanoscale biological objects. TOC Graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=103 SRC="FIGDIR/small/505029v1_ufig1.gif" ALT="Figure 1"> View larger version (21K): org.highwire.dtl.DTLVardef@18e9f99org.highwire.dtl.DTLVardef@900912org.highwire.dtl.DTLVardef@4398baorg.highwire.dtl.DTLVardef@1dff103_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2022

Ratiometric Sensing of Redox Environments Inside Individual Carboxysomes Trapped in Solution

Diffusion of biological nanoparticles in solution impedes our ability to continuously monitor individuals and measure their physical and chemical properties. To overcome this, we previously developed the Interferometric Scattering Anti-Brownian ELectrokinetic (ISABEL) trap, which uses scattering to localize a particle and applies electrokinetic forces which counteract Brownian motion, thus enabling extended observation. Here, we present an improved ISABEL trap that incorporates a near-infrared scatter illumination beam and rapidly interleaves 405 and 488 nm fluorescence excitation reporter beams. With the ISABEL trap, we monitor the internal redox environment of individual carboxysomes labeled with the ratiometric redox reporter roGFP2. Carboxysomes widely vary in scattering contrast (reporting on size) and redox-dependent ratiometric fluorescence. Further, we used redox sensing to explore the chemical kinetics within intact carboxysomes, where bulk measurements may contain unwanted contributions from aggregates or interfering fluorescent proteins. Overall, we demonstrate the ISABEL traps ability to sensitively monitor nanoscale biological objects, enabling new experiments on these systems. TOC Graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=167 SRC="FIGDIR/small/484789v1_ufig1.gif" ALT="Figure 1"> View larger version (45K): org.highwire.dtl.DTLVardef@1fccfacorg.highwire.dtl.DTLVardef@a639b6org.highwire.dtl.DTLVardef@116fff6org.highwire.dtl.DTLVardef@fe95fd_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗