Engineering in Life Sciences最新文献

During the manufacturing of drug substances (DS), endotoxins are a commonly monitored contaminant and checked to ensure the safety and quality of the final product. In this study, we investigated endotoxin removal by commercial anion exchange membrane adsorbers during buffer preparation for an ultrafiltration and diafiltration (UFDF) unit operation as a risk mitigation strategy to meet DS endotoxin specifications. A bracketing design study was used to compare two quaternary amine (Q) modified membranes and one guanidinium functionalized hybrid adsorber for endotoxin removal from spiked buffers representing UFDF start and diafiltration buffer matrices. For the UFDF start buffer, Q-based membrane adsorbers were effective at the removal of endotoxin to the limit of detection in low ionic strength conditions, with one adsorber effective up to 24 mS/cm. Switching to a more complex UFDF diafiltration buffer, Q-based membrane adsorbers were impacted by additional buffer components. The guanidinium-based hybrid membrane adsorber demonstrated endotoxin reduction to the limit of detection from both buffer matrices, showing removal across a wide pH range (4.7–8.3) and conductivity as high as 43 mS/cm. These results demonstrate an operational window for buffers and selected membrane adsorbers to mitigate risk by limiting endotoxin contamination prior to UFDF operations in pharmaceutical applications.

Polydimethylsiloxane (PDMS) is extensively utilized for the recovery of bio-alcohols, but it encounters significant obstacles in volatile organic compounds (VOCs) removal, because of the narrow size for molecules diffusion. In this work, we designed a high-efficiency diffusion channel by introducing phenyl as a spacer into PDMS chains. The monomer divinylbenzene and vinyl-terminated PDMS (vinyl-PDMS) can be chemically crosslinked with thiol-grafted PDMS (thiol-PDMS) based on thiol-ene click reaction. The result shows that the free volume radius (r₃, r₄) has a significant increase after the introduction of divinylbenzene as a spacer, which is beneficial to the transport of phenol diffusion. After a series of optimizations involving the divinylbenzene content, pervaporation (PV) operating temperature, photoinitiator content, and viscosity of vinyl-PDMS, the prepared phenyl-PDMS showed an excellent PV performance for phenol recovery containing 10.9 of separation factor and 3959.66 g m⁻² h⁻¹ of flux as separating 0.1 wt% of phenol/water solution at 70°C. This separation performance is significantly higher than the unmodified PDMS membrane, that is, 2.05 times higher in separation factor and 3.54 times higher in flux. This study provides an effective structure design for the removal of aromatic compounds by enlarging diffusion channels and will make a great contribution to biological medicine and bioengineering.

Circulating tumor cells (CTCs) hold significant promise for cancer diagnosis, prognosis, and treatment monitoring. We previously developed a technique for a single-cell filtering device known as the microcavity array (MCA), specifically designed for the efficient recovery of CTCs from whole blood samples. Efficient enrichment and release of cells from the MCA remains challenging because of cell adhesion that occurs on the MCA surface during the enrichment phase. This study investigated the effects of surface modification with 2-methacryloyloxyethyl phosphorylcholine (MPC) on the recovery efficiency of cancer cell lines from MCA. Scanning electron microscope (SEM) demonstrated reduced cell-substrate interactions, leading to improved recovery efficiency. Comparative analyses showed that the MCA method provided superior recovery efficiency and reduced processing time compared to traditional methods such as density gradient centrifugation (DGC), while maintaining cell viability and proliferative capacity. CTCs were successfully detected in patients with gastric cancer, and short-term cultures were achieved even when fewer than 20 CTCs per milliliter of blood were isolated. These findings emphasize the importance of surface modification for enhancing CTC isolation and the need for optimized culture conditions. The optimized MCA method offers a promising approach for rapid CTC recovery and potential integration with automated systems.

Practical application: The Microcavity array (MCA) is a device specifically designed for efficient recovery of CTCs from whole blood. However cell adhesion on the MCA surface can limit release efficiency. This study demonstrated that surface modification with MPC signigicantly reduces cell-substrate adhesion, improving recovery efficiency while maintaining cell viability and proliferative capacity. Compared to traditional density gradient centrifugation, the MPC-modified MCA offers shorter processing time and better performance. CTCs were successfully detected in gastric cancer, and short-term cultures were achieved even when fewer than 20 CTCs per mL of blood were isolated. The method supports downstearm applications such as cancer cell characterization and treatment monitoring. With potential for integration into automated system, the optimized MCA provides a practical, scalable solution for clinical liquid biopsy and personalized oncology.

Marburg marburgvirus (MARV) is a highly virulent human pathogen with limited therapeutic options. Recombinant MARV glycoprotein (GP) produced in Drosophila Schneider 2 (S2) cells has been extensively investigated as potential vaccine antigen with promising efficacy demonstrated in nonhuman primate models. However, the existing production process for MARV-GP involving static batch cell cultures with limited scalability and process control show lower than desirable yields. Here, we assessed various process intensification strategies in single-use orbital shaken bioreactors (OSBs) or rocking bioreactors (WAVE) and report maximum viable cell concentrations (VCCs) of 31.6 × 10⁶ cells/mL in batch, 69.5 × 10⁶ cells/mL in fed-batch (FB), and up to 210.0 × 10⁶ cells/mL in perfusion mode. By changing from a glucose-only feed to a CellBoost5 feed, MARV-GP yields were increased by over two-fold. Implementation of perfusion cultures achieved a peak MARV-GP concentration of 57.4 mg/L and a 540% higher space-time yield compared to the FB process in the 50 L WAVE system. However, maximum cell-specific productivities were achieved at a VCC of 85 × 10⁶ cells/mL and decreased with increasing cell concentrations. Glycoanalysis revealed a uniform paucimannosidic N-glycan profile, predominantly α-1,6-core-fucosylated Man3F (F(6)M3) structures, across all production modes. Notably, transitioning pH control from CO₂ to phosphoric acid shifted glycan profiles toward higher mannose forms, highlighting the influence of culture conditions on glycosylation.

Sonochemistry has become increasingly important in bioengineering research, and many in vitro and in vivo bioapplications have been developed. Cytotoxicity is always a concern in its implementation. For in vivo treatments and studies, mechanical index (MI) is known to ensure biocompatibility, and even in vitro MI has been used. Because cell characteristics and acoustic phenomena differ in vitro and in vivo, we questioned using MI in vitro. The in vitro cytotoxicity of ultrasound exposure should be investigated to support the development of cutting-edge sonochemistry. In this study, a system for irradiating cultured cells with 1–2 MHz-range ultrasound was developed to demonstrate the invalidity of employing MI alone in vitro. The results showed that cell damage is defined by the MI, ultrasound frequency, and exposure time, which are new indices for quantifying cell damage. Furthermore, cavitation and acoustic streaming are shown to be the main scientific factors that injure cells.