Higher order transient membrane protein structures
Yuxi Zhang,
Hisham Mazal,
Venkata Shiva Mandala,
Gonzalo Perez-Mitta,
Vahid Sandoghdar,
Christoph A. Haselwandter,
Roderick McKinnon
Proceedings of the National Academy of Sciences of the United States of America
122
e2421275121
(2024)
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This study shows that five membrane proteins—three GPCRs, an ion channel, and an enzyme—form self-clusters under natural expression levels in a cardiac-derived cell line. The cluster size distributions imply that these proteins self-oligomerize reversibly through weak interactions. When the concentration of the proteins is increased through heterologous expression, the cluster size distributions approach a critical distribution at which point a phase transition occurs, yielding larger bulk phase clusters. A thermodynamic model like that explaining micellization of amphiphiles and lipid membrane formation accounts for this behavior. We propose that many membrane proteins exist as oligomers that form through weak interactions, which we call higher-order transient structures (HOTS). The key characteristics of HOTS are transience, molecular specificity, and a monotonically decreasing size distribution that may become critical at high concentrations. Because molecular specificity invokes self-recognition through protein sequence and structure, we propose that HOTS are genetically encoded supramolecular units.
Supported Lipid Bilayers as Stochastic Conveyor Belt for Delivery to the Near Field of Nanoscopic Structures
Yazgan Tuna,
Vahid Sandoghdar
Journal of Physical Chemistry C
129
495-499
(2024)
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Placement of nanoscopic entities in the optical near field of a structure such as a plasmonic nanoantenna or the aperture of a scanning near-field optical microscopy (SNOM) remains a nontrivial task, which often requires sophisticated nanofabrication techniques. Here, we show that the fluidity and diffusion of lipid molecules in bilayer geometries can be exploited for facile delivery of nano-objects such as organic dye molecules, semiconductor quantum dots, and gold nanoparticles to the near field of well-defined surface structures. We demonstrate this in three different scenarios with substantial plasmonic enhancement of fluorescence.
Long-Range Three-Dimensional Tracking of Nanoparticles Using Interferometric Scattering Microscopy
Tracking nanoparticle movement is highly desirable in many scientific areas, and various imaging methods have been employed to achieve this goal. Interferometric scattering (iSCAT) microscopy has been particularly successful in combining very high spatial and temporal resolution for tracking small nanoparticles in all three dimensions. However, previous works have been limited to an axial range of only a few hundred nanometers. Here, we present a robust and efficient measurement and analysis strategy for three-dimensional tracking of nanoparticles at high speed and with nanometer precision. After discussing the principle of our approach using synthetic data, we showcase the performance of the method by tracking gold nanoparticles with diameters ranging from 10 to 80 nm in water, demonstrating an axial tracking range from 4 μm for the smallest particles up to over 30 μm for the larger ones. We point out the limitations and robustness of our system across various noise levels and discuss its promise for applications in cell biology and material science, where the three-dimensional motion of nanoparticles in complex media is of interest.
iSCAT microscopy and particle tracking with tailored spatial coherence
Mahdi Mazaheri,
Kiarash Kasaian,
David Albrecht,
Jan Renger,
Tobias Utikal,
Cornelia Holler,
Vahid Sandoghdar
Interferometric scattering (iSCAT) microscopy has demonstrated unparalleled performance among label-free optical methods for detecting and imaging isolated nanoparticles and molecules. However, when imaging complex structures such as biological cells, the superposition of the scattering fields from different locations of the sample leads to a speckle-like background, posing a significant challenge in deciphering fine features. Here, we show that by controlling the spatial coherence of the illumination, one can eliminate the spurious speckle without sacrificing sensitivity. We demonstrate this approach by positioning a rotating diffuser coupled with an adjustable lens and an iris in the illumination path. We report on imaging at a high frame rate of 25 kHz and across a large field of view of 100µm×100µm, while maintaining diffraction-limited resolution. We showcase the advantages of these features by three-dimensional (3D) tracking over 1000 vesicles in a single COS-7 cell and by imaging the dynamics of the endoplasmic reticulum (ER) network. Our approach opens the door to the combination of label-free imaging, sensitive detection, and 3D high-speed tracking using wide-field iSCAT microscopy.
High-Resolution Cryogenic Spectroscopy of Single Molecules in Nanoprinted Crystals
Mohammad Musavinezhad,
Jan Renger,
Johannes Zirkelbach ,
Tobias Utikal,
Claudio U. Hail,
Thomas Basché,
Dimos Poulikakos,
Stephan Götzinger,
Vahid Sandoghdar
We perform laser spectroscopy at liquid helium temperatures (T = 2 K) to investigate single dibenzoterrylene (DBT) molecules doped in anthracene crystals of nanoscopic height fabricated by electrohydrodynamic dripping. Using high-resolution fluorescence excitation spectroscopy, we show that zero-phonon lines of single molecules in printed nanocrystals are nearly as narrow as the Fourier-limited transitions observed for the same guest–host system in the bulk. Moreover, the spectral instabilities are comparable to or less than one line width. By recording super-resolution images of DBT molecules and varying the polarization of the excitation beam, we determine the dimensions of the printed crystals and the orientation of the crystals’ axes. Electrohydrodynamic printing of organic nano- and microcrystals is of interest for a series of applications, where controlled positioning of quantum emitters with narrow optical transitions is desirable.
Measuring Concentration of Nanoparticles in Polydisperse Mixtures Using Interferometric Nanoparticle Tracking Analysis
Anna D. Kashkanova,
David Albrecht,
Michelle Küppers,
Martin Blessing,
Vahid Sandoghdar
Quantitative measurements of nanoparticle concentration in liquid suspensions are in high demand, for example, in the medical and food industries. Conventional methods remain unsatisfactory, especially for polydisperse samples with overlapping size ranges. Recently, we introduced interferometric nanoparticle tracking analysis (iNTA) for high-precision measurement of nanoparticle size and refractive index. Here, we show that by counting the number of trajectories that cross the focal plane, iNTA can measure concentrations of subpopulations in a polydisperse mixture in a quantitative manner and without the need for a calibration sample. We evaluate our method on both monodisperse samples and mixtures of known concentrations. Furthermore, we assess the concentration of SARS-CoV-2 in supernatant samples obtained from infected cells.
An optofluidic antenna for enhancing the sensitivity of single-emitter measurements
Luis Morales-Inostroza,
Julian Folz,
Ralf Kühnemuth,
Suren Felekyan,
Franz Wieser,
Claus A.M. Seidel,
Stephan Götzinger,
Vahid Sandoghdar
Nature Communications
15
2545
(2024)
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Many single-molecule investigations are performed in fluidic environments, e.g., to avoid unwanted consequences of contact with surfaces. Diffusion of molecules in this arrangement limits the observation time and the number of collected photons, thus, compromising studies of processes with fast or slow dynamics. Here, we introduce a planar optofluidic antenna (OFA), which enhances the fluorescence signal from molecules by about 5 times per passage, leads to about 7-fold more frequent returns to the observation volume, and significantly lengthens the diffusion time within one passage. We use single-molecule multi-parameter fluorescence detection (sm-MFD), fluorescence correlation spectroscopy (FCS) and Förster resonance energy transfer (FRET) measurements to characterize our OFAs. The antenna advantages are showcased by examining both the slow (ms) and fast (50μs) dynamics of DNA four-way (Holliday) junctions with real-time resolution. The FRET trajectories provide evidence for the absence of an intermediate conformational state and introduce an upper bound for its lifetime. The ease of implementation and compatibility with various microscopy modalities make OFAs broadly applicable to a diverse range of studies.
Exploring the Physics of Basic Medical Research
Vahid Sandoghdar
Physical Review Letters
132
090001
(2024)
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The 20th century witnessed the emergence of many paradigm-shifting technologies from the physics community, which have revolutionized medical diagnostics and patient care. However, fundamental medical research has been mostly guided by methods from areas such as cell biology, biochemistry, and genetics, with fairly small contributions from physicists. In this Essay, I outline some key phenomena in the human body that are based on physical principles and yet govern our health over a vast range of length and time scales. I advocate that research in life sciences can greatly benefit from the methodology, know-how, and mindset of the physics community and that the pursuit of basic research in medicine is compatible with the mission of physics.<br><br>
invited essay
A paintbrush for delivery of nanoparticles and molecules to live cells with precise spatiotemporal control
Cornelia Holler,
Richard W. Taylor,
Alexandra Schambony,
Leonhard Möckl,
Vahid Sandoghdar
Delivery of very small amounts of reagents to the near-field of cells with micrometer spatial precision and millisecond time resolution is currently out of reach. Here we present μkiss as a micropipette-based scheme for brushing a layer of small molecules and nanoparticles onto the live cell membrane from a subfemtoliter confined volume of a perfusion flow. We characterize our system through both experiments and modeling, and find excellent agreement. We demonstrate several applications that benefit from a controlled brush delivery, such as a direct means to quantify local and long-range membrane mobility and organization as well as dynamical probing of intercellular force signaling.
Contact
Nano-Optics Division Prof. Vahid Sandoghdar
Max Planck Institute for the Science of Light Staudtstr. 2 91058 Erlangen, Germany
