Continuous protein crystallisation platform and process: Case of lysozyme

© 2018 Institution of Chemical Engineers In this work, we designed and built a continuous crystallisation oscillatory flow platform. The lysozyme crystallisation behaviours were investigated at concentrations from 30 to 100 mg/mL, under oscillatory conditions with amplitude (x0) from 10 to 25 mm and frequency (f) from 0.05 to 0.25 Hz in a batch oscillatory flow crystallisation platform. The nucleation rate increased with increase in concentration of initial lysozyme solution, and was also found to increase with increase in shear rate. By learning the thermodynamics and kinetics of lysozyme crystallisation in batch oscillatory flow, the batch crystallisation process was successfully transferred to a continuous oscillatory flow crystallisation process. The equilibrium state of continuous crystallisation reached at residence time 200 min, and the final product crystals shape and size were consistent during the continuous process. This work demonstrates the feasibility of oscillatory flow based platforms for the development of continuous protein crystallisation as for downstream bioseparation

Optimization of vapor diffusion conditions for anti-CD20 crystallization and scale-up to meso batch

© 2019, MDPI AG. All rights reserved. The crystal form is one of the preferred formulations for biotherapeutics, especially thanks to its ability to ensure high stability of the active ingredient. In addition, crystallization allows the recovery of a very pure drug, thus facilitating the manufacturing process. However, in many cases, crystallization is not trivial, and other formulations, such as the concentrate solution, represent the only choice. This is the case of anti-cluster of differentiation 20 (anti-CD20), which is one of the most sold antibodies for therapeutic uses. Here, we propose a set of optimized crystallization conditions for producing anti-CD20 needle-shaped crystals within 24 h in a very reproducible manner with high yield. High crystallization yield was obtained with high reproducibility using both hanging drop vapor diffusion and meso batch, which is a major step forward toward further scaling up the crystallization of anti-CD20. The influence of anti-CD20 storage conditions and the effect of different ions on the crystallization processes were also assessed. The crystal quality and the high yield allowed the first crystallographic investigation on anti-CD20, which positively confirmed the presence of the antibody in the crystals

Optimization of vapor diffusion conditions for anti-CD20 crystallization and scale-up to meso batch

© 2019, MDPI AG. All rights reserved. The crystal form is one of the preferred formulations for biotherapeutics, especially thanks to its ability to ensure high stability of the active ingredient. In addition, crystallization allows the recovery of a very pure drug, thus facilitating the manufacturing process. However, in many cases, crystallization is not trivial, and other formulations, such as the concentrate solution, represent the only choice. This is the case of anti-cluster of differentiation 20 (anti-CD20), which is one of the most sold antibodies for therapeutic uses. Here, we propose a set of optimized crystallization conditions for producing anti-CD20 needle-shaped crystals within 24 h in a very reproducible manner with high yield. High crystallization yield was obtained with high reproducibility using both hanging drop vapor diffusion and meso batch, which is a major step forward toward further scaling up the crystallization of anti-CD20. The influence of anti-CD20 storage conditions and the effect of different ions on the crystallization processes were also assessed. The crystal quality and the high yield allowed the first crystallographic investigation on anti-CD20, which positively confirmed the presence of the antibody in the crystals

In situ Assembly of Raspberry- and Mulberry-like Silica Nanospheres toward Antireflective and Antifogging Coatings

Raspberry- and mulberry-like hierarchically structured silica particulate coatings were fabricated via facile in situ layer-by-layer assembly with monodisperse silica nanoparticles (NPs) of two different sizes followed by calcination. Raspberry-like and mulberry-like silica particulate coatings were achieved when the size ratio of two silica particles was 20/200 and 20/70 nm, respectively. The latter coating exhibited good antireflective property. Its maximum transmittance reached as high as 97%, whereas that of the glass substrate is only 91%. The morphologies of the coatings were observed by scanning electron microscopy and atom force microscopy. The surface properties of these coatings were investigated by measuring their water contact angles and the spreading time of water droplet. The results showed that such hierarchically structured coatings had superhydrophilic and antifogging properties

Synthesis of Raspberry-Like SiO₂–TiO₂ Nanoparticles toward Antireflective and Self-Cleaning Coatings

Silica–titania core–shell nanoparticles of 30, 40, 50, 55, 75, and 110 nm were prepared from tetraethyl orthosilicate (TEOS) and tetraisopropyl titanate (TIPT). After calcination, the amorphous titania shell transformed into anatase nanoparticles, and the silica–titania core–shell nanoparticles became raspberry-like nanoparticles. These nanoparticles were characterized by scanning electron microscopy (SEM), transmission electron microscopy (TEM), thermogravimetric analysis (TGA), X-ray diffraction (XRD), Fourier transform infrared (FT-IR) spectroscopy, and UV–vis spectroscopy. Hierarchically structured antireflective and self-cleaning particulate coatings were fabricated on glass substrates via layer-by-layer (LbL) assembly using silica–titania core–shell nanoparticles and silica nanoparticles as building blocks followed by calcination. The maximum transmittance of coated glass substrates reached as high as ca. 97%, while that of the glass substrates is only ca. 91%. The morphologies of the coatings were observed by SEM and atom force microscopy (AFM). Such hierarchically structured raspberry-like SiO₂–TiO₂ nanoparticle coatings had superhydrophilic and antifogging properties. The coatings also showed photocatalytic activity toward organic pollutants and thus a self-cleaning property

Global Gene Expression Analysis of Cellular Death Mechanisms Induced by Mesoporous Silica Nanoparticle-Based Drug Delivery System

Mesoporous silica nanoparticles (MSNs), as one of the most promising inorganic drug carriers, have attracted ever increasing attention due to their unique structural, physicochemical, and biochemical features. Drug delivery systems (DDSs) based on MSNs could easily escape from endosomes after endocytosis and protect the loaded drugs from bioerosion by stable MSN carriers, efficiently deliver drugs intracellularly in a sustained release way, and consequently kill cancer cells at enhanced efficacy. However, the underlying pathways and mechanisms of cancer cell death induced by MSN-mediated drug delivery have not been well explored. In this study, we introduce gene expression analyses to evaluate the pathways and mechanisms of cancer cell death induced by a MSN-based drug delivery system. Unique changes in gene expressions and gene ontology terms, which were caused only by the MSN-based DDS (DOX-loaded MSNs, DOX@MSNs) but not by free drug doxorubicin (DOX) and/or the carrier MSNs, were discovered and proposed to be responsible for the varied cell death mechanisms, including the greatly enhanced necrosis due to amplified oxidative stress and the apoptosis related with DNA/RNA synthesis and cell cycle inhibitions. By virtue of a certain kind of synergetic biological effect between the drug and the carrier, the DOX@MSNs DDS was found capable of increasing the intracellular levels of reactive oxygen species and triggering the mitochondria-related autophagic lysosome pathway, consequently activating a specific pathway of necrosis, which is different from those by the free drug and the carrier

Relationships between carbon fluxes and environmental factors in a drip-irrigated, film-mulched cotton field in arid region - Fig 5

<p>The response curves of total half-hourly GPP (gross primary production) to mean half-hourly R_n (net solar radiation) (a), T_air (air temperature) (b), and VPD (vapor pressure deficit) (c) at the 95% confidence interval.</p

Circulating Fibrocytes Are Increased in Neonates with Bronchopulmonary Dysplasia - Fig 3

<p>Correlation between circulating fibrocyte counts and right ventricular systolic pressure (A) (Pearson correlation coefficient; r = −0.62; p < 0.01) or oxygen saturation (B) (Pearson correlation coefficient; r = −0.62; p < 0.01).</p

LAI comparison for each of the growth stages.

<p>LAI comparison for each of the growth stages.</p

Energy closure ratio comparsions of mulched cotton in the Wulanwusu and non-mulched vegetations in other areas.

<p>Energy closure ratio comparsions of mulched cotton in the Wulanwusu and non-mulched vegetations in other areas.</p