A Basic Method for Formulating mRNA-Lipid Nanoparticle Vaccines in the Lab.

During recent years, RNA therapeutics have begun to make a substantial impact in the clinic, with the approval of the siRNA-based therapeutic Patisiran in 2018, and of the two mRNA SARS-CoV-2 vaccines, BNT162b2 and mRNA-1273 in 2021. A key to the success of these therapeutics lies in the lipid-based delivery system. The therapeutic RNAs are encapsulated in lipid nanoparticles (LNPs), which protect against enzymatic degradation and efficiently deliver the RNA across the cell membrane into the cytosol.

Design and Synthesis of Circular RNA Expression Vectors.

The recent success of the synthetic mRNA-based anti-COVID-19 vaccines has demonstrated the broad potential of the mRNA platform for applications in medicine, thanks to the combined efforts of a small community that has vastly improved key determinants such as design and formulation of synthetic mRNA during the past three decades. However, the cost of production and sensitivity to enzymatic degradation are still limiting the broader application of synthetic mRNA for therapeutic applications. The increased interest in mRNA-based technologies has spurred a renaissance for circular RNA (circRNA), as the lack of free 5' and 3' ends substantially increases resistance against enzymatic degradation in biological systems and does not require expensive cap analogs, as translation is controlled by an Internal Ribosome Entry Site (IRES) sequence.

Dissecting cell membrane tension dynamics and its effect on Piezo1-mediated cellular mechanosensitivity using force-controlled nanopipettes.

The dynamics of cellular membrane tension and its role in mechanosensing, which is the ability of cells to respond to physical stimuli, remain incompletely understood, mainly due to the lack of appropriate tools. Here, we report a force-controlled nanopipette-based method that combines fluidic force microscopy with fluorescence imaging for precise manipulation of the cellular membrane tension while monitoring the impact on single-cell mechanosensitivity. The force-controlled nanopipette enables control of the indentation force imposed on the cell cortex as well as of the aspiration pressure applied to the plasma membrane.

An Approach for the Real-Time Quantification of Cytosolic Protein-Protein Interactions in Living Cells.

In recent years, cell-based assays have been frequently used in molecular interaction analysis. Cell-based assays complement traditional biochemical and biophysical methods, as they allow for molecular interaction analysis, mode of action studies, and even drug screening processes to be performed under physiologically relevant conditions. In most cellular assays, biomolecules are usually labeled to achieve specificity.

Investigating Complex Samples with Molograms of Low-Affinity Binders.

In vitro diagnostics relies on the quantification of minute amounts of a specific biomolecule, called biomarker, from a biological sample. The majority of clinically relevant biomarkers for conditions beyond infectious diseases are detected by means of binding assays, where target biomarkers bind to a solid phase and are detected by biochemical or physical means. Nonspecifically bound biomolecules, the main source of variation in such assays, need to be washed away in a laborious process, restricting the development of widespread point-of-care diagnostics.

Ultra Stable Molecular Sensors by Submicron Referencing and Why They Should Be Interrogated by Optical Diffraction-Part II. Experimental Demonstration.

Label-free optical biosensors are an invaluable tool for molecular interaction analysis. Over the past 30 years, refractometric biosensors and, in particular, surface plasmon resonance have matured to the standard of this field despite a significant cross reactivity to environmental and experimental noise sources. In this paper, we demonstrate that sensors that apply the spatial affinity lock-in principle (part I) and perform readout by diffraction overcome the drawbacks of established refractometric biosensors.

Quantification of Molecular Interactions in Living Cells in Real Time using a Membrane Protein Nanopattern.

Molecular processes within cells have traditionally been studied with biochemical methods due to their high degree of specificity and ease of use. In recent years, cell-based assays have gained more and more popularity since they facilitate the extraction of mode of action, phenotypic, and toxicity information. However, to provide specificity, cellular assays rely heavily on biomolecular labels and tags while label-free cell-based assays only offer holistic information about a bulk property of the investigated cells.

Image reversal reactive immersion lithography improves the detection limit of focal molography: erratum.

Some minor issues were discovered after publication of Opt Lett. 43, 5801 (2018) and are corrected here. They do not change the main message of the paper.

An analytical method to control the surface density and stability of DNA-gold nanoparticles for an optimized biosensor.

DNA functionalized gold nanoparticles (DNA-AuNPs) have shown great potential for biosensing as they combine the excellent optical properties of gold nanoparticles and the molecular recognition function of DNA. Since the DNA density determines the assay performance and the stability of the conjugate, a precise control of the surface density of DNA-AuNP is crucial for an optimized biosensor. Here we report a simple assay for quantifying multiple unlabeled DNAs on AuNPs.

Image reversal reactive immersion lithography improves the detection limit of focal molography.

Focal molography is a label-free optical biosensing method that relies on a coherent pattern of binding sites for biomolecular interaction analysis. Reactive immersion lithography (RIL) is central to the patterning of molographic chips but has potential for improvements. Here, we show that applying the idea of image reversal to RIL enables the fabrication of coherent binding patterns of increased quality (i.

A guide towards long-term functional electrodes interfacing neuronal tissue.

Implantable electronics address therapeutical needs of patients with electrical signaling dysfunctions such as heart problems, neurological disorders or hearing impairments. While standard electronics are rigid, planar and made of hard materials, their surrounding biological tissues are soft, wet and constantly in motion. These intrinsic differences in mechanical and chemical properties cause physiological responses that constitute a fundamental challenge to create functional long-term interfaces.

Lipid Nanoparticle Assisted mRNA Delivery for Potent Cancer Immunotherapy.

The induction of a strong cytotoxic T cell response is an important prerequisite for successful immunotherapy against many viral diseases and tumors. Nucleotide vaccines, including mRNA vaccines with their intracellular antigen synthesis, have been shown to be potent activators of a cytotoxic immune response. The intracellular delivery of mRNA vaccines to the cytosol of antigen presenting immune cells is still not sufficiently well understood.

mRNA vaccine delivery using lipid nanoparticles.

mRNA vaccines elicit a potent immune response including antibodies and cytotoxic T cells. mRNA vaccines are currently evaluated in clinical trials for cancer immunotherapy applications, but also have great potential as prophylactic vaccines. Efficient delivery of mRNA vaccines will be key for their success and translation to the clinic.