Publications & technical resources

Explore how DHO technology is facilitating scientific discovery

Showing
publications
Thank you! Your submission has been received!
Oops! Something went wrong while submitting the form.
Education
Materials science
Image refocusing
Darkfield
Brightfield
Polarization imaging
Simultaneous multicolor
FISH & smFISH
Custom order
Tetrapod
Deep focus
Single helix
Double helix
Super resolution
Multiplexed imaging
High-throughput screening
High-content analysis
Inspection
Data visualization
Data analysis
Image reconstruction
Emitter localization
OEM integrations
Offline inspection
Online inspection
Spatial omics
Volumetric imaging
Nuclear biology
Drug discovery
Bacterial biology
FRET & smFRET
HILO
TIRF
Widefield
PAINT
PALM
STORM
Computer vision
Multicolor
Light sheet
Optical engineering
Immunotherapy
Chemical engineering
CLEM
Biophysics
3D particle tracking
Physical chemistry
Neuroscience
Proteomics
SPINDLE
Phase mask
Cell biology
Variable-angle illumination
3DTRAX
3D SMLM
Drug delivery
Two photon microscopy
Genomics
Environmental remediation
AI & ML
May 1, 2024
|
Biomedical Optics Express
Tyler Nelson, Sofía Vargas-Hernández, Margareth Freire, Siyang Cheng, and Anna-Karin Gustavsson
Single-molecule super-resolution imaging is instrumental in investigating cellular architecture and organization at the nanoscale. Achieving precise 3D nanometric localization when imaging structures throughout mammalian cells, which can be multiple microns thick, requires careful selection of the illumination scheme in order to optimize the fluorescence signal to background ratio (SBR). Thus, an optical platform that combines different wide-field illumination schemes for target-specific SBR optimization would facilitate more precise 3D nanoscale studies of a wide range of cellular structures. Here, we demonstrate a versatile multimodal illumination platform that integrates the sectioning and background reduction capabilities of light sheet illumination with homogeneous, flat-field epi- and TIRF illumination. Using primarily commercially available parts, we combine the fast and convenient switching between illumination modalities with point spread function engineering to enable 3D single-molecule super-resolution imaging throughout mammalian cells. For targets directly at the coverslip, the homogenous intensity profile and excellent sectioning of our flat-field TIRF illumination scheme improves single-molecule data quality by providing low fluorescence background and uniform fluorophore blinking kinetics, fluorescence signal, and localization precision across the entire field of view. The increased contrast achieved with LS illumination, when compared with epi-illumination, makes this illumination modality an excellent alternative when imaging targets that extend throughout the cell. We validate our microscopy platform for improved 3D super-resolution imaging by two-color imaging of paxillin – a protein located in the focal adhesion complex – and actin in human osteosarcoma cells.
View publication
Feb 21, 2024
|
bioRxiv
Pratim Chowdhury, Xiaoli Wang, Julia F. Love, Sofia Vargas-Hernandez, Yuya Nakatani, Sandra L. Grimm, Dereck Mezquita, Frank M. Mason, Elisabeth D. Martinez, Cristian Coarfa, Cheryl L. Walker, Anna-Karin Gustavsson, and Ruhee Dere
Centrosomes play a fundamental role in nucleating and organizing microtubules in the cell and are vital for faithful chromosome segregation and maintenance of genomic stability. Loss of structural or functional integrity of centrosomes causes genomic instability and is a driver of oncogenesis. The lysine demethylase 4A (KDM4A) is an epigenetic ‘eraser’ of chromatin methyl marks, which we show also localizes to the centrosome with single molecule resolution. We additionally discovered KDM4A demethylase enzymatic activity is required to maintain centrosome homeostasis, and is required for centrosome integrity, a new functionality unlinked to altered expression of genes regulating centrosome number. We find rather, that KDM4A interacts with both mother and daughter centriolar proteins to localize to the centrosome in all stages of mitosis. Loss of KDM4A results in supernumerary centrosomes and accrual of chromosome segregation errors including chromatin bridges and micronuclei, markers of genomic instability. In summary, these data highlight a novel role for an epigenetic ‘eraser’ regulating centrosome integrity, mitotic fidelity, and genomic stability at the centrosome.
View publication
Oct 17, 2023
|
Biophysical Journal
Stephen L. Upton, Jian W. Tay, Daniel K. Schwartz, and Marcelo C. Sousa
The β-barrel assembly machinery (BAM) complex is responsible for inserting outer membrane proteins (OMPs) into the Escherichia coli outer membrane. The SecYEG translocon inserts inner membrane proteins into the inner membrane and translocates both soluble proteins and nascent OMPs into the periplasm. Recent reports describe Sec possibly playing a direct role in OMP biogenesis through interactions with the soluble polypeptide transport-associated (POTRA) domains of BamA (the central OMP component of BAM). Here we probe the diffusion behavior of these protein complexes using photoactivatable super-resolution localization microscopy and single-particle tracking in live E. coli cells of BAM and SecYEG components BamA and SecE and compare them to other outer and inner membrane proteins. To accurately measure trajectories on the highly curved cell surface, three-dimensional tracking was performed using double-helix point-spread function microscopy. All proteins tested exhibit two diffusive modes characterized by “slow” and “fast” diffusion coefficients. We implement a diffusion coefficient analysis as a function of the measurement lag time to separate positional uncertainty from true mobility. The resulting true diffusion coefficients of the slow and fast modes showed a complete immobility of full-length BamA constructs in the time frame of the experiment, whereas the OMP OmpLA displayed a slow diffusion consistent with the high viscosity of the outer membrane. The periplasmic POTRA domains of BamA were found to anchor BAM to other cellular structures and render it immobile. However, deletion of individual distal POTRA domains resulted in increased mobility, suggesting that these domains are required for the full set of cellular interactions. SecE diffusion was much slower than that of the inner membrane protein PgpB and was more like OMPs and BamA. Strikingly, SecE diffused faster upon POTRA domain deletion. These results are consistent with the existence of a BAM-SecYEG trans-periplasmic assembly in live E. coli cells.
View publication
Sep 28, 2023
|
Nature Structural & Molecular Biology
S. Basu, O. Shukron, D. Hall, P. Parutto, A. Ponjavic, D. Shah, W. Boucher, D. Lando, W. Zhang, N. Reynolds, L. H. Sober, A. Jartseva, R. Ragheb, X. Ma, J. Cramard, R. Floyd, J. Balmer, T. A. Drury, A. R. Carr, L.-M. Needham, A. Aubert, G. Communie, K. Gor, M. Steindel, L. Morey, E. Blanco, T. Bartke, L. Di Croce, I. Berger, C. Schaffitzel, S. F. Lee, T. J. Stevens, D. Klenerman, B. D. Hendrich, and D. Holcman
To understand how the nucleosome remodeling and deacetylase (NuRD) complex regulates enhancers and enhancer–promoter interactions, we have developed an approach to segment and extract key biophysical parameters from live-cell three-dimensional single-molecule trajectories. Unexpectedly, this has revealed that NuRD binds to chromatin for minutes, decompacts chromatin structure and increases enhancer dynamics. We also uncovered a rare fast-diffusing state of enhancers and found that NuRD restricts the time spent in this state. Hi-C and Cut&Run experiments revealed that NuRD modulates enhancer–promoter interactions in active chromatin, allowing them to contact each other over longer distances. Furthermore, NuRD leads to a marked redistribution of CTCF and, in particular, cohesin. We propose that NuRD promotes a decondensed chromatin environment, where enhancers and promoters can contact each other over longer distances, and where the resetting of enhancer–promoter interactions brought about by the fast decondensed chromatin motions is reduced, leading to more stable, long-lived enhancer–promoter relationships.
View publication
Apr 25, 2023
|
Journal of Biomedical Optics
Jingjing Gao, Pengwei Wang, Wenwen Li, Xuyu Zhang, Chunyuan Song, Zhentao Liu, Shensheng Han, and Honglin Liu
Significance: Double-helix point spread function (DH-PSF) microscopy has been developed for three-dimensional (3D) localization and imaging at super-resolution but usually in environments with no or weak scattering. To date, super-resolution imaging through turbid media has not been reported. Aim: We aim to explore the potential of DH-PSF microscopy in the imaging and localization of targets in scattering environments for improved 3D localization accuracy and imaging quality. Approach: The conventional DH-PSF method was modified to accommodate the scanning strategy combined with a deconvolution algorithm. The localization of a fluorescent microsphere is determined by the center of the corresponding double spot, and the image is reconstructed from the scanned data by deconvoluting the DH-PSF. Results: The resolution, i.e., the localization accuracy, was calibrated to 13 nm in the transverse plane and 51 nm in the axial direction. Penetration thickness could reach an optical thickness (OT) of 5. Proof-of-concept imaging and the 3D localization of fluorescent microspheres through an eggshell membrane and an inner epidermal membrane of an onion are presented to demonstrate the super-resolution and optical sectioning capabilities. Conclusions: Modified DH-PSF microscopy can image and localize targets buried in scattering media using super-resolution. Combining fluorescent dyes, nanoparticles, and quantum dots, among other fluorescent probes, the proposed method may provide a simple solution for visualizing deeper and clearer in/through scattering media, making in situ super-resolution microscopy possible for various demanding applications.
View publication

No results found

Please try different keywords

Showing
webinars
Thank you! Your submission has been received!
Oops! Something went wrong while submitting the form.

No results found

Please try different keywords

Showing
notes
Thank you! Your submission has been received!
Oops! Something went wrong while submitting the form.

No results found

Please try different keywords

Ready to learn more?