Jongyoon Han

Perfusion bioreactor systems enable intensified bioprocessing with high cell densities and volumetric productivities in monoclonal antibody (mAb) manufacturing. However, their widespread adoption remains constrained by the high consumption of chemically defined cell culture media. In this study, we introduce a multi‐stage electrokinetic waste separation process that regenerates spent media, recovering up to 87.5% of the spent‐medium volume for reuse. Using a mock‐perfusion model, the regeneration‐recycle processes were shown to sustain high‐density Chinese hamster ovary cell growth, compared with spent media recycling without regeneration. Integration into a self‐recycling perfusion bioreactor (350 mL working volume) operating with 75% regenerated media sustained the target viable cell density and antibody titer, with limited reductions to cell‐specific growth rate (~31%) and cell‐specific productivity (~9%), supporting the feasibility of high‐ratio media recycling. Techno‐economic modeling of large‐scale scenarios revealed that media regeneration could offer a path to $15/g COGs, thereby increasing the accessibility of mAbs. Techno‐economic modeling of hypothetical large‐scale manufacturing scenarios indicated that the experimentally demonstrated process could reduce cost of goods (COG) by approximately 9%, and identified a theoretical pathway toward ~$15/g COG contingent upon multi‐cycle regeneration processes that could be developed in the future. These results support regenerative perfusion as a promising process‐intensification strategy that, with further development, may be adaptable to other mammalian cell culture platforms where media cost and sustainability are critical bottlenecks.

Lentiviral vectors (LVVs) are widely used as gene‐delivery vehicles in cell and gene therapy, enabling stable integration of transgenes into target cells. However, efficient clarification of LVV harvest remains a major bottleneck in large‐scale manufacturing, as conventional depth filtration often results in significant product loss. Here, we introduce a fouling‐free microfluidic cell clarification (MCC) system that enables high‐efficiency LVV recovery. By precisely controlling the harvest flow rate and adjusting the recirculated cell density by adding buffer, we established a quantitative relationship between clarification efficiency and throughput. Under optimized conditions, the MCC achieved > 95% product recovery while maintaining > 95% cell removal efficiency. The MCC process closely followed the theoretical recovery model assuming negligible product loss within the device. Across three independent batch operations, apparent average LVV recovery remained substantially higher than that of a benchmark process employing a cellulose depth filter‐based process. Scalability tests demonstrated consistent performance at flow rates up to 1.5 L/h without loss of clarification efficiency, demonstrating its potential for large‐scale biomanufacturing. Beyond LVVs, the MCC platform is broadly applicable to cell‐based modalities that secrete therapeutic products, offering a continuous, low‐shear, and fouling‐free alternative to conventional filtration processes.

A 25-layer spiral microfluidic device significantly reduces cell biomass from high-turbidity feedstock using inertial focusing to perform primary clarification and harvest adeno-associated virus (AAV) vectors at 20 mL min ⁻¹ with 85% vector recovery.

Nuclear magnetic resonance relaxometry measurements have a diverse array of applications in medicine, biology, and chemistry, and they are valuable for monitoring cell cultures, diagnosing diseases, and detecting defects in pharmaceutical products. However, relaxometry workflows typically require laborious procedures and large sample volumes, which are both slow and destructive. This makes it difficult to use relaxometry in applications such as drug screening and cell culture development, where it has significant potential to be useful. In this work, we describe and demonstrate a low-cost benchtop system that can perform relaxometry measurements on 16 wells of a 96-well culture plate in one shot, enabling rapid, nondestructive scanning of multiple groups without any sample preparation. We demonstrate the application of this multiwell scanner for iron measurement and parallel measurements.

A spiral inertial microfluidic cell retention device enables high-density HEK293 perfusion bioprocessing for intensified lentiviral vector production, yielding functional titers up to 10 ⁸ TU mL ⁻¹ through the device's harvest stream.

Background: Clinical application of mesenchymal stromal cells (MSCs) for cartilage regeneration has been hampered by their poor retention of chondrogenic potential during in vitro culture, which has led to highly variable cartilage repair outcomes. Consequently, there is an urgent need for a reliable assay to predict MSC chondrogenic potential during cell manufacturing. Methods: In this study, we developed a nondestructive MSCs iron flux monitoring methodology using spent culture media, utilizing micromagnetic resonance relaxometry (µMRR). We demonstrated that the dynamics of iron uptake and release by MSCs in culture can be reliably inferred from iron concentration changes in culture medium, with an unprecedented temporal resolution (<1 hour). Results: Analysis of 3 MSC donors across 6 independent culture batches revealed a significant correlation between iron homeostasis (iron flux during culture) and chondrogenic differentiation outcomes, while a departure from iron homeostasis (significant iron uptake and accumulation) was correlated with impaired chondrogenesis. By contrast, cell proliferation, although essential for manufacturing to achieve sufficient cell numbers, did not reliably correlate with chondrogenic capacity. Furthermore, ascorbic acid supplementation during culture, which is known to promote MSCs proliferation and chondrogenic quality, regulated iron homeostasis by limiting iron flux. Conclusion: Our findings identify iron homeostasis as a potential chondrogenic-associated critical quality attribute of MSCs. This rapid, nondestructive monitoring strategy offers a promising approach to improve manufacturing efficiency and consistency by providing real-time insight into cellular iron flux. In addition, the methodology here impacts broader iron biology by providing a time-resolved iron flux measurement that is not currently available.

Acute myocardial infarction (AMI), a leading cause of death globally, triggers complex inflammatory responses critical to patient outcomes. However, rapid tools for profiling immune responses at the single‐cell level are lacking. The integrated Single‐cell Enzyme and Antigen Quantification (iSEAQ) system addresses this gap by enabling high‐throughput, single‐cell analysis of immune cell activity using just 20 μL of blood. This novel tool processes live CD66b and CD3 cells to quantify the secretion of Granzyme B, Neutrophil Elastase, and CD31 within minutes. Longitudinal studies on nine AMI patients revealed that CD66b ⁺ cells are major contributors (up to 95%) to key inflammatory enzymes, including the unexpected secretion of Granzyme B. iSEAQ achieves unparalleled sensitivity (0.4 fg/cell) and predictive accuracy (>90%) for patient profiling across AMI onset, treatment, and discharge. This innovation provides clinicians with a rapid, precise method to monitor immune responses, unveiling new insights into AMI inflammation and therapy.

Conventional iron determination methods, such as colorimetric assays, often lack sensitivity at low iron levels and are susceptible to interference from complex compositions in serum and plasma. In this study, we developed a rapid, sensitive, and accurate method for iron determination in 1 min utilizing an inexpensive benchtop 0.5-T micromagnetic resonance relaxometry (μMRR) system with acidification treatment of samples (pH < 1). The method yielded a highly linear calibration curve (R² > 0.999) between the transverse relaxation rate R₂ and iron concentration from 0.5 to 1000 μM, with a limit of detection (LOD) of 0.25 μM and a minimal assay volume of 5 μL. The accuracy of this method was validated by inductively coupled plasma mass spectrometry (ICP-MS) across a diverse range of biological samples. This μMRR-based approach offers a rapid and convenient alternative for serum and plasma iron measurements, which can substantially reduce clinical diagnostic time and support real-time iron monitoring for patients.