Biology subjects

Shahrokhtash, A.

Publications and source records attributed to Shahrokhtash, A..

3 recordsLinked to original sources

bioRxiv2025

Nanoscale precise stamping of biomolecule patterns using DNA origami

Understanding the importance of ligand patterning in biological processes requires precise control over molecular positioning and spacing. While DNA origami structures offer nanoscale precision in biomolecule arrangement, their biological applications are limited by challenges related to their structural stability, scalability, and surface area. Here, we present a straightforward and rapid DNA origami stamping technique for transferring nanoscale oligonucleotide patterns onto surfaces, visualized using DNA-PAINT super-resolution microscopy to quantitatively assess the stamping efficiency and precision across different stamp types. Unlike traditional top-down methods that require specialized equipment, our technique provides an accessible, self-assembled platform for surface patterning, with versatility across various substrates via modifiable pattern-transfer oligonucleotides. We demonstrate reliable, efficient, and precise pattern transfer at single-molecule resolution, enabling new opportunities to study distance-dependent biological processes, including receptor activation, multivalent binding, and enzymatic cascades across broader spatial scales and different detection techniques. The use of the passivated surface limits non-specific interactions with unpatterned areas and enables control over the interaction between the biological target and the patterned biomolecules. Our method advances surface patterning by combining DNA nanotechnology with single-molecule imaging techniques, expanding access to cost-effective analytical approaches and potentially enabling multiplexed detection and live measurements.

bioengineering↗

bioRxiv2025

Serial intravital microscopy reveals temporal dynamics of autoreactive germinal centers in the spleen

The spleen plays a key role in clearing blood-borne infections and is involved in autoimmune and hematological disorders. It undergoes extensive remodeling during inflammation and immune reactions, but its localization in the peritoneal cavity has hampered studies of these dynamic changes. Here, we establish and validate a protocol for serial 2-photon microscopy of the murine spleen to capture dynamic processes in the living animal. As a proof-of-principle, we elucidate the expansion and contraction of autoreactive germinal centers (GCs) induced by epicutaneous application of the small-molecule TLR7 agonist resiquimod (R848). Leveraging a biocompatible abdominal imaging window, intravital labeling techniques, and fluorescent reporters, we follow GCs up to 180 {micro}m below the capsule for more than 2 weeks by tracking follicular dendritic cell (FDC) networks. This was accomplished without appreciable perturbation of normal physiology, paving the way for a deeper understanding of the biology of the spleen and its associated disease states. HighlightAn abdominal imaging window allowing the study of dynamic processes in the spleen of live mice over the course of several weeks.

immunology↗

bioRxiv2023

Microplate Format Protein Nanopatterning for High-Throughput Screening of Cellular Microenvironments

An advanced protein nanopatterned cell culture platform is engineered to emulate the extracellular matrixs complexity, enabling precise nanoscale biomolecule copatterning to mimic environments analogous to native tissue for cellular assays. Nanopatterns fabricated through sparse colloidal lithography, with 100 nm to 800 nm features in separate wells, are seamlessly integrated into standard microplate formats (96-well/384-well). Robust patterns are built from fully PEGylated, passivated thin glass coverslips optimized for minimal nonspecific interactions. Biotin-avidin binding and click chemistry to ensure the accurate and robust localization of bioligands. The transparent, metal-free substrates are free of topographical interference, rendering them ideal for diverse fluorescence microscopy techniques encompassing single-molecule TIRFM and extensive high-throughput imaging. The structural stability of these nanopatterns persists beyond a year in storage and long-term in cell culture conditions, endorsing their application for prolonged experimental studies and potential for widespread academic and industrial use. The platform has been demonstrated for nanopatterning an array of biomolecules, from small molecules to proteins, DNA, and extracellular matrix components, instrumental for studying cell signaling. Experiments with C2C12 cells demonstrated the exceptional specificity of the nanopatterned microplates, with nonspecific adhesion remaining below 2% and the platforms ability to elicit size-dependent cellular reactions when interfaced with nanopatterned fibronectin.

bioengineering↗