Biology subjects

Chizhik, A. I.

Publications and source records attributed to Chizhik, A. I..

4 recordsLinked to original sources

bioRxiv2025

Plasmonic Nanocavity-Based Method for Measuring Electric Charges in Solution

Electric charges play a fundamental role in shaping the structure, function, and interactions of biomolecules, yet precisely measuring these charges at the single-molecule level remains a significant technical challenge. Here, we introduce a novel experimental methodology that utilizes plasmonic nanocavities to quantify molecular electric charges in solution with high sensitivity. Our approach exploits an externally applied electric field to induce the spatial redistribution of charged molecules confined within a planar metallic nanocavity, while simultaneously leveraging nanocavity-induced fluorescence lifetime modulation as a highly sensitive readout. We demonstrate the feasibility of this method through proof-of-concept experiments, where we measure the fluorescence lifetimes of positively and negatively charged fluorescent dye molecules as a function of the applied electric field across the cavity. The experimental results are validated through a rigorous theoretical framework, incorporating statistical thermodynamics and electrodynamic modeling to accurately describe the observed data. The proposed method offers a calibration-free, experimentally simple, and rapid alternative to existing charge measurement techniques, opening new avenues for precise quantification of molecular electric charges down to the single-molecule level.

biophysics↗

bioRxiv2022

Metal-Induced Energy Transfer (MIET) for Live-Cell Imaging with Fluorescent Proteins

Metal-Induced Energy Transfer (MIET) imaging is an easy-to-implement super-resolution modality that achieves nanometer resolution along the optical axis of a microscope. Although its capability in numerous biological and biophysical studies has been demonstrated, its implementation for live-cell imaging with fluorescent proteins is still lacking. Here, we present its applicability and capabilities for live-cell imaging with fluorescent proteins in diverse cell types (adult human stem cells, human osteo-sarcoma cells, and Dictyostelium discoideum cells), and with various fluorescent proteins (GFP, mScarlet, RFP, YPet). We show that MIET imaging achieves nanometer axial mapping of living cellular and sub-cellular components across multiple timescales, from a few milliseconds to hours, with negligible phototoxic effects.

biophysics↗

bioRxiv2021

Isotropic Three-Dimensional Dual-Color Super-Resolution Microscopy with Metal-Induced Energy Transfer

Over the last two decades, super-resolution microscopy has seen a tremendous development in speed and resolution, but for most of its methods, there exists a remarkable gap between lateral and axial resolution. Similar to conventional optical microscopy, the axial resolution is by a factor three to five worse than the lateral resolution. One recently developed method to close this gap is metal-induced energy transfer (MIET) imaging which achieves an axial resolution down to nanometers. It exploits the distance dependent quenching of fluorescence when a fluorescent molecule is brought close to a metal surface. In the present manuscript, we combine the extreme axial resolution of MIET imaging with the extraordinary lateral resolution of single-molecule localization microscopy, in particular with direct stochastic optical reconstruction microscopy (dSTORM). This combination allows us to achieve isotropic three-dimensional super-resolution imaging of sub-cellular structures. Moreover, we employed spectral demixing for implementing dualcolor MIET-dSTORM that allows us to image and co-localize, in three dimensions, two different cellular structures simultaneously.

biophysics↗

bioRxiv2021

Mapping Activity-Dependent Quasi-Stationary States of Mitochondrial Membranes with Graphene-Induced Energy Transfer Imaging

Graphene-induced energy transfer (GIET) was recently introduced for the precise localization of fluorescent molecules along the optical axis of a microscope. GIET is based on near-field energy transfer from an optically excited fluorophore to a single sheet of graphene. As a proof-of-concept, we demonstrated its potential by determining the distance between the two leaflets of supported lipid bilayers (SLBs) with sub-nanometer accuracy. Here, we use GIET imaging for three-dimensional reconstruction of the mitochondrial membrane architecture. We map two quasi-stationary states of the inner and outer mitochondrial membranes before and during adenosine tri-phosphate (ATP) synthesis. We trigger the ATP synthesis state in vitro by activating mitochondria with precursor molecules. Our results demonstrate that the inner membrane (IM) approaches the outer membrane (OM) while the outer membrane (OM) does not show a measurable change in average axial position upon activation. As a result, the inter-membrane space (IM-OM distance) is reduced by [~]2 nm upon activation of the mitochondria. This direct experimental observation of the subtle dynamics of mitochondrial membranes and the change in inter-membrane distance induced by ATP synthesis is relevant for our understanding of the physical functioning of mitochondria.

biophysics↗