Delicate Molecules Research Articles

One-Dimensional “Ghost Imaging” in Electron Microscopy of Inelastically Scattered Electrons

Entanglement and correlation are at the basis of quantum mechanics and have been used in optics to create a framework for “ghost imaging”. We propose that a similar scheme can be used in an electron microscope to exploit the correlation of electrons with the coincident detection of collective mode excitations in a sample. In this way, an image of the sample can be formed on an electron camera even if electrons never illuminated the region of interest directly. This concept, which can be regarded as the inverse of photon-induced near-field electron microscopy, can be used to probe delicate molecules with a resolution that is beyond the wavelength of the collective mode.

Tailoring Lipid-Based Drug Delivery Nanosystems by Synchrotron Small Angle X-ray Scattering.

Thanks to specific physico-chemical properties, drug delivery systems based on nanoparticles have proven to effectively transport delicate molecules for therapeutic purposes, protecting them from degradation, increasing their stability in the blood circulation and allowing to convey and release the transported substances in specific areas of the body. Nanoparticles obtained from biopolymers for applications in medicine and pharmaceutics have become particularly popular in recent years due to the enormous research effort in the field of vaccines to respond to the pandemic emergency. Among the various types of biopolymers used to produce nanoparticles for therapeutics, lipids have characteristics that make them biocompatible, with low toxicity and ease of clearance. They can be synthesized by designing their characteristics according to the foreseen administration path, or to the target of the transported drug. The analytical methods mostly used to evaluate the characteristics of lipid nanosytems for drug delivery involve studying their effects on cells, in vitro and in vivo. Although it is often considered a "niche technique" for research in the bio-related sciences, Small Angle X-ray Scattering (SAXS) is a versatile tool to study the structure of nanosystems based on lipids, both ex situ and in situ. Therefore, it allows to evaluate both the effect of the different synthesis parameters and of the exposure of lipid nanoparticles to physiological conditions, which is of fundamental importance to design efficient drug delivery systems. In this mini-review, we will report some recent examples of characterization and design of nanoparticles based on lipids, where SAXS has been a fundamental step both to guide the synthesis of nanomaterials with tailored characteristics, and to understand the interaction between nanomaterials and cells.

Techniques and Applications of Protein Mass Spectrometry

Protein mass spectrometry demands that proteins in solution or solid form be ionised in the gas phase before being injected and accelerated in an electric or magnetic field for analysis. Electrospray Ionisation (ESI) and matrix-assisted laser desorption/ionization are the two most used techniques for ionising proteins. Electrospray allows delicate molecules to be ionised whole, sometimes retaining non-covalent connections, since the ions are generated from proteins in solution. In MALDI, proteins are placed in a matrix that is typically solid, and ions are generated by laser light pulses. Electrospray generates more multiply-charged ions than MALDI, allowing for higher mass protein measurement and better fragmentation for identification, whereas MALDI is faster and less susceptible to contamination, buffers, and additives.

Repairing the heart: State-of the art delivery strategies for biological therapeutics.

Myocardial infarction (MI) is one of the leading causes of mortality worldwide. It is caused by an acute imbalance between oxygen supply and demand in the myocardium, usually caused by an obstruction in the coronary arteries. The conventional therapy is based on the application of (a combination of) anti-thrombotics, reperfusion strategies to open the occluded artery, stents and bypass surgery. However, numerous patients cannot fully recover after these interventions. In this context, new therapeutic methods are explored. Three decades ago, the first biologicals were tested to improve cardiac regeneration. Angiogenic proteins gained popularity as potential therapeutics. This is not straightforward as proteins are delicate molecules that in order to have a reasonably long time of activity need to be stabilized and released in a controlled fashion requiring advanced delivery systems. To ensure long-term expression, DNA vectors-encoding for therapeutic proteins have been developed. Here, the nuclear membrane proved to be a formidable barrier for efficient expression. Moreover, the development of delivery systems that can ensure entry in the target cell, and also correct intracellular trafficking towards the nucleus are essential. The recent introduction of mRNA as a therapeutic entity has provided an attractive intermediate: prolonged but transient expression from a cytoplasmic site of action. However, protection of the sensitive mRNA and correct delivery within the cell remains a challenge. This review focuses on the application of synthetic delivery systems that target the myocardium to stimulate cardiac repair using proteins, DNA or RNA.

Surface modification for patterned cell growth on substrates with pronounced topographies using sacrificial photoresist and parylene-C peel-off

A range of methods including soft lithography are available for patterning protein layers for cell adhesion on quasi-planar substrates. Suitably structured, these layers favor the geometrically constrained, controlled growth of cells and the development of cellular extensions on them. For this purpose, the ability to control the shape and dimension of cell-adhesive areas with high precision is crucial. For more advanced studies of cell interactions, the surface modification or functionalization of substrates with complex topographies is desirable. This paper describes a simple technique allowing to produce surface modification patterns using delicate molecules such as laminin on substrates exhibiting pronounced topographies with recessed and protruding microstructures. The technique is based on the combination of sacrificial photoresist structures with a connected parylene-C layer. This layer locally adheres to the substrate wherever the substrate needs to be protected against the surface modification. After surface modification, the parylene-C layer is peeled off. Patterns comprising arbitrary networks of modified and unmodified substrate areas can thus be realized. We demonstrate the technique with the guided growth of neuron-like PC12 cells on networks of laminin lines on substrates structured with micropillars and microwells.

Lipase immobilization for catalytic applications obtained using fumed silica deposited with MAPLE technique

Lipases are enzymes used for catalyzing reactions of acylglycerides in biodiesel production from lipids, where enzyme immobilization on a substrate is required.Silica nanoparticles in different morphologies and configurations are currently used in conjunction with biological molecules for drug delivery and catalysis applications, but up to date their use for triglycerides has been limited by the large size of long-chain lipid molecules.Matrix assisted pulsed laser evaporation (MAPLE), a laser deposition technique using a frozen solution/suspension as a target, is widely used for deposition of biomaterials and other delicate molecules. We have carried out a MAPLE deposition starting from a frozen mixture containing fumed silica and lipase in water. Deposition parameters were chosen in order to increase surface roughness and to promote the formation of complex structures. Both the target (a frozen thickened mixture of nanoparticles/catalyst in water) and the deposition configuration (a small target to substrate distance) are unusual and have been adopted in order to increase surface contact of catalyst and to facilitate access to long-chain molecules. The resulting innovative film morphology (fumed silica/lipase cluster level aggregation) and the lipase functionality (for catalytic biodiesel production) have been studied by FESEM, FTIR and transesterification tests.

Cold cathode field emitter array on a quadrupole mass spectrometer: Route to miniaturization

We have improved the quadrupole mass spectrometer by substituting the conventional hot filament electron source by a field emitter array (FEA). Elimination of the hot filament avoids a number of common problems, including thermal cracking of delicate molecules, outgassing of the filament itself and nearby components, high power requirements for the filament, large size, stray light, stray magnetic fields, contamination by thoria and tungsten, and a long warm-up time. The advantages are clearest for portable applications where power requirements dominate. Here, the power savings are not just in eliminating the filament supply, but more important in reducing the largest component of the system, the vacuum pump. This comes about because the filament is the primary gas load and because chemical reactions taking place on it require fast pumping to keep the products from interfering with the spectra. Comparison between hot filament and cold cathode FEA ionization is made using a quadrupole mass spectrometer fitted with both electron sources, independently controlled. The FEA advantage is strongest when the ultrahigh vacuum system is throttled to a low pumping speed, mimicking a portable system with a small pump. FEAs also enable miniaturization and a corresponding decrease in pump size. Moreover, with miniaturization, shorter mean free paths and consequently higher working pressures can be tolerated further decreasing pump requirements.