To investigate the potential of two flow imaging microscopy (FIM) techniques (Micro-Flow Imaging (MFI) and FlowCAM) to determine total cell concentration and cell viability. B-lineage acute... Show moreTo investigate the potential of two flow imaging microscopy (FIM) techniques (Micro-Flow Imaging (MFI) and FlowCAM) to determine total cell concentration and cell viability. B-lineage acute lymphoblastic leukemia (B-ALL) cells of 2 different donors were exposed to ambient conditions. Samples were taken at different days and measured with MFI, FlowCAM, hemocytometry and automated cell counting. Dead and live cells from a fresh B-ALL cell suspension were fractionated by flow cytometry in order to derive software filters based on morphological parameters of separate cell populations with MFI and FlowCAM. The filter sets were used to assess cell viability in the measured samples. All techniques gave fairly similar cell concentration values over the whole incubation period. MFI showed to be superior with respect to precision, whereas FlowCAM provided particle images with a higher resolution. Moreover, both FIM methods were able to provide similar results for cell viability as the conventional methods (hemocytometry and automated cell counting). FIM-based methods may be advantageous over conventional cell methods for determining total cell concentration and cell viability, as FIM measures much larger sample volumes, does not require labeling, is less laborious and provides images of individual cells. PURPOSE METHODS RESULTS CONCLUSION Show less
A key enabling step in leveraging the properties of nanoparticles (NPs) is to explore new, simple, controllable, and scalable nanotechnologies for their syntheses. Among “wet” methods, cathodic... Show moreA key enabling step in leveraging the properties of nanoparticles (NPs) is to explore new, simple, controllable, and scalable nanotechnologies for their syntheses. Among “wet” methods, cathodic corrosion has been used to synthesize catalytic aggregates with some control over their size and preferential faceting. Here, we report on a modification of the cathodic corrosion method for producing a range of nonaggregated nanocrystals (Pt, Pd, Au, Ag, Cu, Rh, Ir, and Ni) and nanoalloys (Pt50Au50, Pd50Au50, and AgxAu100–x) with potential for scaling up the production rate. The method employs poly(vinylpyrrolidone) (PVP) as a stabilizer in an electrolyte solution containing nonreducible cations (Na+, Ca2+), and cathodic corrosion of the corresponding wires takes place in the electrolyte under ultrasonication. The ultrasonication not only promotes particle–PVP interactions (enhancing NP dispersion and diluting locally high NP concentration) but also increases the production rate by a factor of ca. 5. Further increase in the production rate can be achieved through parallelization of electrodes to construct comb electrodes. With respect to applications, carbon-supported Pt NPs prepared by the new method exhibit catalytic activity and durability for methanol oxidation comparable or better than the commercial benchmark catalyst. A variety of AgxAu100–x nanoalloys are characterized by ultraviolet–visible absorption spectroscopy and high-resolution transmission electron microscopy. The protocol for NP synthesis by cathodic corrosion should be a step toward its further use in academic research as well as in its practical upscaling. Show less
The aim of this study was to develop a supercritical carbon dioxide (scCO2) spray process to coat solid protein particles with a hydrophilic polymer. The final purpose is to manufacture drug... Show moreThe aim of this study was to develop a supercritical carbon dioxide (scCO2) spray process to coat solid protein particles with a hydrophilic polymer. The final purpose is to manufacture drug particles exhibiting controlled release behaviour in patients. Lysozyme microparticles (about 20 μm) were suspended in a vessel into which a dextran sulphate (DS) solution was dispersed by scCO2 via a nozzle. Upon interaction with the droplets, DS was deposited onto or mixed with suspended lysozyme particles. Particles of about 100 μm were obtained. The zeta-potential analysis and elemental analysis indicated that the top layer of the particles consisted of both lysozyme and DS. Some of the produced particulate materials showed retarded lysozyme release when exposed to water or phosphate buffered saline, holding promise for future production of controlled drug delivery systems for therapeutic proteins. Show less
Complex injectable formulations, such as protein therapeutics, controlled release systems and cell therapy products, are gaining a paramount position in the therapy of many life-threatening and... Show moreComplex injectable formulations, such as protein therapeutics, controlled release systems and cell therapy products, are gaining a paramount position in the therapy of many life-threatening and chronic diseases. Most of these products have in common that sub-visible particles (SVP), i.e., particulate matter in the size range of about 1 – 100 µm, are critical quality attributes. Most protein therapeutics are liquid or freeze-dried formulations in which the presence of SVP is unwanted; many injectable controlled release systems are based on particulate drug delivery systems in the sub-visible size range; and cell concentration and viability are important characteristics of cell therapy products. With the continuous improvement of existing and emerging particle analysis techniques, the potentials of these tools in addressing current characterization challenges in the field of complex injectable formulations have to be investigated. Therefore, the aim of this thesis was to develop methods, based on a set of state-of-the-art particle analysis techniques, for characterization of pharmaceutically relevant sub-visible particles and to study the value of these methods in the characterization of complex injectable formulations. Show less
The purpose of this study was to explore the potential of flow imaging microscopy to measure particle size and agglomeration of poly(lactic-co-glycolic acid) (PLGA) microparticles. The particle... Show moreThe purpose of this study was to explore the potential of flow imaging microscopy to measure particle size and agglomeration of poly(lactic-co-glycolic acid) (PLGA) microparticles. The particle size distribution of pharmaceutical PLGA microparticle products is routinely determined with laser diffraction. In our study, we performed a unique side-by-side comparison between MFI 5100 (flow imaging microscopy) and Mastersizer 2000 (laser diffraction) for the particle size analysis of two commercial PLGA microparticle products, i.e., Risperdal Consta and Sandostatin LAR. Both techniques gave similar results regarding the number and volume percentage of the main particle population (28–220 μm for Risperdal Consta; 16–124 μm for Sandostatin LAR). MFI additionally detected a ‘fines’ population (<28 μm for Risperdal Consta; <16 μm for Sandostatin LAR), which was overlooked by Mastersizer. Moreover, MFI was able to split the main population into ‘monospheres’ and ‘agglomerates’ based on particle morphology, and count the number of particles in each sub-population. Finally, we presented how MFI can be applied in process development of risperidone PLGA microparticles and to monitor the physical stability of Sandostatin LAR. These case studies showed that MFI provides insight into the effect of different process steps on the number, size and morphology of fines, monospheres and agglomerates as well as the extent of microparticle agglomeration after reconstitution. This can be particularly important for the suspendability, injectability and release kinetics of PLGA microparticles. Show less
This study addressed the effect of contact sliding during stirring of a monoclonal antibody solution onprotein aggregation, in particular, in the nanometer and micrometer size range. An overhead... Show moreThis study addressed the effect of contact sliding during stirring of a monoclonal antibody solution onprotein aggregation, in particular, in the nanometer and micrometer size range. An overhead stirring setupwas designed in which the presence and magnitude of the contact between the stir bar and thecontainer could be manipulated. A solution of 0.1 mg/mL of a monoclonal antibody (IgG) in phosphatebuffered saline was stirred at 300 rpm at room temperature. At different time points, samples were takenand analyzed by nanoparticle tracking analysis, flow imaging microscopy, and size-exclusion chromatography.In contrast to nonecontact-stirred and unstirred samples, the contact-stirred sample containedseveral-fold more particles and showed a significant loss of monomer. No increase in oligomer contentwas detected. The number of particles formed was proportional to the contact area and the magnitude ofthe normal pressure between the stir bar and the glass container. Extrinsic 9-(2,2-dicyanovinyl) julolidinefluorescence indicated a conformational change for contact-stirred protein samples. Presence ofpolysorbate 20 inhibited the formation of micron-sized aggregates. We suggest a model in whichabrasion of the potentially destabilized, adsorbed protein leads to aggregation and renewal of the surfacefor adsorption of a fresh protein layer. Show less
Sediq, A.S.; Nejadnik, M.R.; El Bialy, I.; Witkamp, G.J.; Jiskoot, W. 2015
