Development of generic metabolic Raman calibration models using solution titration in aqueous phase and data augmentation for in-line cell culture analysis.

This study presents a novel approach for developing generic metabolic Raman calibration models for in-line cell culture analysis using glucose and lactate stock solution titration in an aqueous phase and data augmentation techniques. First, a successful set-up of the titration method was achieved by adding glucose or lactate solution at several different constant rates into the aqueous phase of a bench-top bioreactor. Subsequently, the in-line glucose and lactate concentration were calculated and interpolated based on the rate of glucose and lactate addition, enabling data augmentation and enhancing the robustness of the metabolic calibration model. Nine different combinations of spectra pretreatment, wavenumber range selection, and number of latent variables were evaluated and optimized using aqueous titration data as training set and a historical cell culture data set as validation and prediction set. Finally, Raman spectroscopy data collected from 11 historical cell culture batches (spanning four culture modes and scales ranging from 3 to 200 L) were utilized to predict the corresponding glucose and lactate values. The results demonstrated a high prediction accuracy, with an average root mean square errors of prediction of 0.65 g/L for glucose, and 0.48 g/L for lactate. This innovative method establishes a generic metabolic calibration model, and its applicability can be extended to other metabolites, reducing the cost of deploying real-time cell culture monitoring using Raman spectroscopy in bioprocesses.

Further accelerating biologics development from DNA to IND: the journey from COVID-19 to non-COVID-19 programs.

The Coronavirus Disease (COVID-19) pandemic has spurred adoption of revolutionary initiatives by regulatory agencies and pharmaceutical industry worldwide to deliver therapeutic COVID-19 antibodies to patients at unprecedented speed. Among these, timeline of chemistry, manufacturing and control (CMC), which involves process development and manufacturing activities critical for the assurance of product quality and consistency before first-in-human clinical trials, was greatly reduced from typically 12-15 months (using clonal materials) to approximately 3 months (using non-clonal materials) in multiple cases. In this perspective, we briefly review the acceleration approaches published for therapeutic COVID-19 antibodies and subsequently discuss the applicability of these approaches to achieve investigational new drug (IND) timelines of ≤10 months in over 60 COVID-19 and non-COVID-19 programs performed at WuXi Biologics. We are of the view that, with demonstrated product quality and consistency, innovative approaches used for COVID-19 can be widely applied in all disease areas for greater speed to clinic.

Advancing injection force modeling and viscosity-dependent injectability evaluation for prefilled syringes.

The development of PFS requires a detailed understanding of the forces occurring during the drug administration process and patient's capability. This research describes an advanced mathematic injection force model that consisting hydrodynamic force and friction force. The hydrodynamic force follows the basic law of Hagen-Poiseuille but refines the modeling approach by delving into specific properties of drug viscosity (Newtonian and Shear-thinning) and syringe shape constant, while the friction force was accounted from empty barrel injection force. Additionally, we take actual temperature of injection into consideration, providing more accurate predication. The results show that the derivation of the needle dimension constant and the rheological behavior of the protein solutions are critical parameters. Also, the counter pressure generated by the tissue has been considered in actual administration to address the issue of the inaccuracies of current injection force evaluation preformed in air, especially when the viscosity of the injected drug solution is below 9.0 cP (injecting with 1 mL L PFS staked with 29G ½ inch needle). Human factor studies on patients' capability against medication viscosity filled the gap in design space of PFS drug product and available viscosity data in very early phase.

Nanoparticle technology for mRNA: Delivery strategy, clinical application and developmental landscape.

The mRNA vaccine, a groundbreaking advancement in the field of immunology, has garnered international recognition by being awarded the prestigious Nobel Prize, which has emerged as a promising prophylactic and therapeutic modality for various diseases, especially in cancer, rare disease, and infectious disease such as COVID-19, wherein successful mRNA treatment can be achieved by improving the stability of mRNA and introducing a safe and effective delivery system. Nanotechnology-based delivery systems, such as lipid nanoparticles, lipoplexes, polyplexes, lipid-polymer hybrid nanoparticles and others, have attracted great interest and have been explored for mRNA delivery. Nanoscale platforms can protect mRNA from extracellular degradation while promoting endosome escape after endocytosis, hence improving the efficacy. This review provides an overview of diverse nanoplatforms utilized for mRNA delivery in preclinical and clinical stages, including formulation, preparation process, transfection efficiency, and administration route. Furthermore, the market situation and prospects of mRNA vaccines are discussed here.

Intensified perfusion culture (IPC) reduced recombinant protein fragmentation.

Mammalian cells remain the mainstay of biological production host. In industry, cultivating and harvest strategies are sorted in batch mode (e.g., batch, fed-batch, concentrated fed-batch and intensified fed-batch) and continuous mode (e.g., perfusion). To retrieve greater productivity and better product quality, especially for the sensitive products prone to fragmentation, culture modes with various modifications are innovated (e.g., intensified perfusion culture [IPC]). In our study, we demonstrated that the fragmentation of Fc-fusion product (Molecule A) is time-dependent in traditional fed-batch (TFB) culture. The fragmentation proportion increased from 3.8% to 12.4% for Clone A, 0.8% to 1.7% for Clone B and 0.9% to 2.0% for Clone C from Day 10 to Day 14. By applying a novel bioprocess, IPC, which allows continuous feeding of the fresh medium and constant removal of the spent medium without bleeding cells to maintain a defined constant viable cell density, the fragmentation was reduced to 0.3% while the productivity was increased from 2.96 g/L to 15.51 g/L for Clone A. To validate whether the fragmentation reduction is product-sensitive, plasmids carrying the DNA sequences of two other Fc-fusion molecules (Molecule B and Molecule C) were transfected into the host. The results showed consistent fragmentation reducing effect by using IPC. Furthermore, the cultivation scale was expanded to 50 L and 1000 L. A minimum fragmentation level below 0.1% was observed for Molecule C. Our study revealed the capability of IPC in reducing Fc-fusion protein fragmentation and the reproducibility when scaling up while maintaining high productivity.

In Situ Regulating Cobalt/Iron Oxide-Oxyhydroxide Exchange by Dynamic Iron Incorporation for Robust Oxygen Evolution at Large Current Density.

The key dilemma for green hydrogen production via electrocatalytic water splitting is the high overpotential required for anodic oxygen evolution reaction (OER). Co/Fe-based materials show superior catalytic OER activity to noble metal-based catalysts, but still lag far behind the state-of-the-art Ni/Fe-based catalysts probably due to undesirable side segregation of FeOOH with poor conductivity and unsatisfied structural durability under large current density. Here, a robust and durable OER catalyst affording current densities of 500 and 1000 mA cm-2 at extremely low overpotentials of 290 and 304 mV in base is reported. This catalyst evolves from amorphous bimetallic FeOOH/Co(OH)2 heterostructure microsheet arrays fabricated by a facile mechanical stirring strategy. Especially, in situ X-ray photoelectron spectroscopy (XPS) and Raman analysis decipher the rapid reconstruction of FeOOH/Co(OH)2 into dynamically stable Co1-x Fex OOH active phase through in situ iron incorporation into CoOOH, which perform as the real active sites accelerating the rate-determining step supported by density functional theory calculations. By coupling with MoNi4 /MoO2 cathode, the self-assembled alkaline electrolyzer can deliver 500 mA cm-2 at a low cell voltage of 1.613 V, better than commercial IrO2 (+) ||Pt/C(-) and most of reported transition metal-based electrolyzers. This work provides a feasible strategy for the exploration and design of industrial water-splitting catalysts for large-scale green hydrogen production.

Developing an ultra-intensified fed-batch cell culture process with greatly improved performance and productivity.

Intensified fed-batch (IFB), a popular cell culture intensification strategy, has been widely used for productivity improvement through high density inoculation followed by fed-batch cultivation. However, such an intensification strategy may counterproductively induce rapidly progressing cell apoptosis and difficult-to-sustain productivity. To improve culture performance, we developed a novel cell culture process intermittent-perfusion fed-batch (IPFB) which incorporates one single or multiple cycles of intermittent perfusion during an IFB process for better sustained cellular and metabolic behaviors and notably improved productivity. Unlike continuous perfusion or other semi-continuous processes such as hybrid perfusion fed-batch with only early-stage perfusion, IPFB applies limited times of intermittent perfusion in the mid-to-late stage of production and still inherits bolus feedings on nonperfusion days as in a fed-batch culture. Compared to IFB, an average titer increase of ~45% was obtained in eight recombinant CHO cell lines studied. Beyond IPFB, ultra-intensified IPFB (UI-IPFB) was designed with a markedly elevated seeding density of 20-80 × 106 cell/mL, achieved through the conventional alternating tangential flow filtration (ATF) perfusion expansion followed with a cell culture concentration step using the same ATF system. With UI-IPFB, up to ~6 folds of traditional fed-batch and ~3 folds of IFB productivity were achieved. Furthermore, the application grounded in these two novel processes showed broad-based feasibility in multiple cell lines and products of interest, and was proven to be effective in cost of goods reduction and readily scalable to a larger scale in existing facilities.

A Review of Recent FDA-Approved Biologic-Device Combination Products.

With the remarkably strong growth of the biopharmaceutical market, an increasing demand for self-administration and rising competitions attract substantial interest to the biologic-device combination products. The ease-of-use of biologic-device combination products can minimize dosing error, improve patient compliance and add value to the life-cycle management of biological products. As listed in the purple book issued by the U.S. Food and Drug Administration (FDA), a total of 98 brand biologic-device combination products have been approved with Biologic License Application from January 2000 to August 2023, where this review mainly focused on 63 products containing neither insulin nor vaccine. Prefilled syringes (PFS) and autoinjectors are the most widely adopted devices, whereas innovative modifications like needle safety guard and dual-chamber design and novel devices like on-body injector also emerged as promising presentations. All 16 insulin products employ pen injectors, while all 19 vaccine products are delivered by a PFS. This review provides a systematic summary of FDA-approved biologic-device combination products regarding their device configurations, routes of administration, formulations, instructions for use, etc. In addition, challenges and opportunities associated with biologic-device compatibility, regulatory complexity, and smart connected devices are also discussed. It is believed that evolving technologies will definitely move the boundaries of biologic-device combination product development even further.

Green Synthesis and Morphological Evolution for Bi2Te3 Nanosystems via a PVP-Assisted Hydrothermal Method.

Bi2Te3 has been extensively used because of its excellent thermoelectric properties at room temperature. Here, 230-420 nm of Bi2Te3 hexagonal nanosheets has been successfully synthesized via a "green" method by using ethylene glycol solution and applying polyvinyl pyrrolidone (PVP) as a surfactant. In addition, factors influencing morphological evolution are discussed in detail in this study. Among these parameters, the reaction temperature, molar mass of NaOH, different surfactants, and reaction duration are considered as the most essential. The results show that the existence of PVP is vital to the formation of a plate-like morphology. The reaction temperature and alkaline surroundings played essential roles in the formation of Bi2Te3 single crystals. By spark plasma sintering, the Bi2Te3 hexagonal nanosheets were hot pressed into solid-state samples. We also studied the transport properties of solid-state samples. The electrical conductivity σ was 18.5 × 103 Sm-1 to 28.69 × 103 Sm-1, and the Seebeck coefficient S was -90.4 to -113.3 µVK-1 over a temperature range of 300-550 K. In conclusion, the observation above could serve as a catalyst for future exploration into photocatalysis, solar cells, nonlinear optics, thermoelectric generators, and ultraviolet selective photodetectors of Bi2Te3 nanosheet-based photodetectors.

Low-Voltage Solution-Processed Zinc-Doped CuI Thin Film Transistors with NOR Logic and Artificial Synaptic Function.

Low-voltage Zn-doped CuI thin film transistors (TFTs) gated by chitosan dielectric were fabricated at a low temperature. The Zn-doped CuI TFT exhibited a more superior on/off current ratio than CuI TFT due to the substitution or supplementation of copper vacancies by Zn ions. The Zn-doped CuI films were characterized by scanning electron microscope, X-ray diffraction, and X-ray photoelectron spectroscopy. The Zn-doped CuI TFTs exhibited an on/off current ratio of 1.58 × 104, a subthreshold swing of 70 mV/decade, and a field effect mobility of 0.40 cm2V-1s-1, demonstrating good operational stability. Due to the electric-double-layer (EDL) effect and high specific capacitance (17.3 µF/cm2) of chitosan gate dielectric, Zn-doped CuI TFT operates at a voltage below -2 V. The threshold voltage is -0.2 V. In particular, we have prepared Zn-doped CuI TFTs with two in-plane gates and NOR logic operation is implemented on such TFTs. In addition, using the ion relaxation effect and EDL effect of chitosan film, a simple pain neuron simulation is realized on such a p-type TFTs for the first time through the bottom gate to regulate the carrier transport of the channel. This p-type device has promising applications in low-cost electronic devices, complementary electronic circuit, and biosensors.

Correlation between the Protein Pharmaceutical Surface Activity and Interfacial Stability.

The interaction of protein drugs with the air-liquid interface plays a crucial role in the overall stability in aqueous formulations, particularly when the adsorbed proteins are subjected to the surface flow. Nonionic surfactants are usually added into the formulation solutions to address this issue. A diversity of studies have been focused on the usage of surfactants, the stability mechanism of surfactants, or seeking new pharmaceutical surfactants. However, the real protagonist, the basic properties of protein drugs, was neglected, which may play a vital role in the stability of protein drugs. Herein, we aim to clarify the correlation between the surface behavior of proteins and the interfacial stability. A force tensiometer is used to track the surface tension reduction and the competition between surfactants and proteins at the surface. We find that the surface behaviors of proteins vary with storage temperature and protein types including monoclonal antibodies (mAb), bispecific monoclonal antibodies (BsAb), and antibody-drug conjugates (ADCs). Especially for the protein stored at 5 °C, the surface activity of proteins is better than that of surfactants. It indicates that the ability of proteins to adsorb at the interface should not be ignored compared to surfactants. The significant difference in the interfacial stability of protein pharmaceuticals formulated in the same buffer and excipients as well as the surfactants with the same concentration further confirms the interfacial adsorption capacity of proteins that should not be ignored. These findings provide a new angle and valuable insights into the correlation between the surface activity of the proteins and interfacial stability, which may pave the way for future preformulation studies on therapeutic proteins and broaden the thoughts of formulation development.

Modulating and optimizing Pluronic F-68 concentrations and feeding for intensified perfusion Chinese hamster ovary cell cultures.

Perfusion culture is often performed with micro-sparger to fulfill the high oxygen demand from the densified cells. Protective additive Pluronic F-68 (PF-68) is widely used to mitigate the adverse effect in cell viability from micro-sparging. In this study, different PF-68 retention ratio in alternating tangential filtration (ATF) columns was found to be crucial for cell performance of different perfusion culture modes. The PF-68 in the perfusion medium was found retained inside the bioreactor when exchanged through ATF hollow fibers with a small pore size (50 kD). The accumulated PF-68 could provide sufficient protection for cells under micro-sparging. On the other hand, with large-pore-size (0.2 µm) hollow fibers, PF-68 could pass through the ATF filtration membranes with little retention, and consequently led to compromised cell growth. To overcome the defect, a PF-68 feeding strategy was designed and successfully verified on promoting cell growth with different Chinese hamster ovary (CHO) cell lines. With PF-68 feeding, enhancements were observed in both viable cell densities (20%-30%) and productivity (~30%). A threshold PF-68 concentration of 5 g/L for high-density cell culture (up to 100 × 106 cells/mL) was also proposed and verified. The additional PF-68 feeding was not observed to affect product qualities. By designing the PF-68 concentration of perfusion medium to or higher than the threshold level, a similar cell growth enhancement was also achieved. This study systematically investigated the protecting role of PF-68 in intensified CHO cell cultures, shedding a light on the optimization of perfusion cultures through the control of protective additives.

Acetate accumulation and regulation by process parameters control in Chinese hamster ovary cell culture.

Chinese hamster ovary (CHO) cells represent a group of predominantly used mammalian hosts for producing recombinant therapeutic proteins. Known for their rapid proliferation rates, CHO cells undergo aerobic glycolysis that is characterized by fast glucose consumption, that ultimately gives rise to a group of small-molecule organic acids. However, only the function of lactate has been extensively studied in CHO cell culture. In this study, we observed the accumulation of acetate from the late exponential phase to harvest day, potentially contributing to the pH decline in late culture stage regardless of lactate consumption. In addition, we evaluated the acidification of the fresh media and the cell culture suspension, and the data revealed that acetate presented a lower acidification capacity compared to lactate and exhibited limited inhibitory effect on cells with less than 20 mM supplemented in the media. This study also explored the ways to control acetate accumulation in CHO cell culture by manipulating the process parameters such as temperature, glucose, and pH control. The positive correlation between the specific glucose consumption rate and acetate generation rate provides evidence of the endogenous acetate generation from overflow metabolism. Reducing these parameters (temperature, glucose consumption) and HCl-controlled low pH ultimately suppress acetate build-up. In addition, the specific acetate generation rate and relevant glucose consumption rate are found to be a metabolic trait associated with specific cell lines. Taken together, the results presented in these experiments provide a means to advance industrial CHO cell culture process control and development.

Rapidly accelerated development of neutralizing COVID-19 antibodies by reducing cell line and CMC development timelines.

Since the Coronavirus Disease 2019 (COVID-19) outbreak, unconventional cell line development (CLD) strategies have been taken to enable development of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-neutralizing antibodies at expedited speed. We previously reported a novel chemistry, manufacturing, and control (CMC) workflow and demonstrated a much-shortened timeline of 3-6 months from DNA to investigational new drug (IND) application. Hereafter, we have incorporated this CMC strategy for many SARS-CoV-2-neutralizing antibody programs at WuXi Biologics. In this paper, we summarize the accelerated development of a total of seven antibody programs, some of which have received emergency use authorization  approval in less than 2 years. Stable pools generated under good manufacturing practice (GMP) conditions consistently exhibited similar productivity and product quality at different scales and batches, enabling rapid initiation of phase I clinical trials. Clones with comparable product quality as parental pools were subsequently screened and selected for late-stage development and manufacturing. Moreover, a preliminary stability study plan was devised to greatly reduce the time required for final clone determination and next-generation sequencing-based viral testing was implemented to support rapid conditional release of the master cell bank for GMP production. The successful execution of these COVID-19 programs relies on our robust, fit for purpose, and continuously improving CLD platform. The speed achieved for pandemic-related biologics development may innovate typical biologics development timelines and become a new standard in the industry.

Competition of Photo-Excitation and Photo-Desorption Induced Positive and Negative Photoconductivity Switch in Te Nanowires.

The photocurrent in tellurium nanowire (Te NW) exhibits a subtle influence by many extrinsic factors. Herein, we fabricate Te NW devices and explore their photoresponse properties in detail. It is observed that the current increases greatly at low environmental relative humidity (RH) under light illumination, demonstrating an evident positive photoconductivity (PPC). However, the photocurrent reduces at high RH, yielding a typical negative photoconductivity (NPC). In addition, when exposed to a proper relative humidity, Te NW devices show PPC immediately and then transfer to NPC gradually under illumination, exhibiting the RH sensitive PPC/NPC switch. It is proposed that the competition between photo-excitation and photo-desorption is responsible for this subtle switch of PPC/NPC. On the one hand, the adsorbed water molecules on the surface of Te nanowires, acting as electron acceptors, lead to an increase of conductance, exhibiting the PPC phenomenon. On the other hand, the photo-desorption of water molecules from the surface results in a decreased carrier concentration in the Te nanowires, yielding the NPC phenomenon. The in-depth understanding of such charge transfer processes between the absorbed water molecules and Te nanowires provides an effective route to modulate the carrier densities and control the PPC/NPC switch, which will accelerate the design and application of novel optoelectronic nanodevices.

Ferroelectric-gated MoSe2photodetectors with high photoresponsivity.

Ferroelectric transistors with semiconductors as the channel material and ferroelectrics as the gate insulator have potential applications in nanoelectronics. We report in-situ modulation of optoelectronic properties of MoSe2thin flakes on ferroelectric 0.7PbMg1/3Nb2/3O3-0.3PbTiO3(PMN-PT). Under the excitation of 638 nm laser, the photoresponsivity can be greatly boosted to 59.8 A W-1and the detectivity to 3.2 × 1010Jones, with the improvement rates of about 1500% and 450%, respectively. These results suggest hybrid structure photodetector of two-dimensional layered material and ferroelectric has great application prospects in photoelectric detector.

Productivity and quality improvement for a symmetric bispecific antibody through the application of intensified perfusion cell culture.

Background: Aggregation, fragmentation, and low yield are issues frequently found during the cell culture process of bispecific antibodies (bsAbs), whose inherent complexity likely plays a role in causing these issues. Methods: In this study, we made a head-to-head comparison between fed-batch cell culture and intensified perfusion cell culture with a symmetric bsAb case. Results: In comparison with the fed-batch culture, a 6.6-fold improvement in integrated viable cell density and a 10.9-fold improvement in volumetric productivity were achieved with the intensified perfusion mode. In addition, a significant decrease in aggregation and fragmentation was observed with the intensified perfusion cell culture. Furthermore, product homogeneity was improved, which was reflected by the increased percentage of capillary isoelectric focusing main group. The quality improvement with intensified perfusion cell culture can be attributed to the shortened product retention in the bioreactor. Conclusions: These findings suggest that intensified perfusion cell culture could be a better choice than traditional fed-batch especially for complex molecules like bsAbs. As this is a single case report, future studies on other cases are needed to further confirm the general applicability of this strategy.

Approaching intrinsic dynamics of MXenes hybrid hydrogel for 3D printed multimodal intelligent devices with ultrahigh superelasticity and temperature sensitivity.

Hydrogels are investigated broadly in flexible sensors which have been applied into wearable electronics. However, further application of hydrogels is restricted by the ambiguity of the sensing mechanisms, and the multi-functionalization of flexible sensing systems based on hydrogels in terms of cost, difficulty in integration, and device fabrication remains a challenge, obstructing the specific application scenarios. Herein, cost-effective, structure-specialized and scenario-applicable 3D printing of direct ink writing (DIW) technology fabricated two-dimensional (2D) transition metal carbides (MXenes) bonded hydrogel sensor with excellent strain and temperature sensing performance is developed. Gauge factor (GF) of 5.7 (0 - 191% strain) and high temperature sensitivity (-5.27% °C-1) within wide working range (0 - 80 °C) can be achieved. In particular, the corresponding mechanisms are clarified based on finite element analysis and the first use of in situ temperature-dependent Raman technology for hydrogels, and the printed sensor can realize precise temperature indication of shape memory solar array hinge.

Optimization of hydrogen-ion storage performance of tungsten trioxide nanowires by niobium doping.

The transport and storage of ions within solid state structures is a fundamental limitation for fabricate more advanced electrochemical energy storage, memristor, and electrochromic devices. Crystallographic shear structure can be induced in the tungsten bronze structures composed of corner-sharing WO6octahedra by the addition of edge-sharing NbO6octahedra, which might provide more storage sites and more convenient transport channels for external ions such as hydrogen ions and alkali metal ions. Here, we show that Nb2O5·15WO3nanowires (Nb/W = 0.008) with long length-diameter ratio, smooth surface, and uniform diameter have been successfully synthesized by a simple hydrothermal method. The Nb2O5·15WO3nanowires do exhibit more advantages over h-WO3nanowires in electrochemical hydrogen ion storage such as smaller polarization, larger capacity (71 mAh g-1, at 10C, 1C = 100 mA g-1), better cycle performance (remain at 99% of the initial capacity after 200 cycles at 100C) and faster H+ions diffusion kinetics. It might be the crystallographic shear structure induced by Nb doping that does result in the marked improvement in the hydrogen-ion storage performance of WO3. Therefore, complex niobium tungsten oxide nanowires might offer great promise for the next generation of electrochemical energy and information storage devices.

Optimized process operations reduce product retention and column clogging in ATF-based perfusion cell cultures.

Product retention in hollow fibers is a common issue in ATF-based cell culture system. In this study, the effects of four major process factors on product (therapeutic antibody/recombinant protein) retention were investigated using Chinese hamster ovary cell. Hollow fibers made of polysulfone presented a product retention rate from 15% ± 8 to 43% ± 18% higher than those made of polyether sulfone varying with specific processes. Higher harvest flowrate and ATF exchange rate increased product retention by 13% ± 10% and up to 31% ± 13%, respectively. Hollow fibers with larger pore sizes (0.65 µm) appeared to have increased product retention by 38% ± 7% compared with smaller ones (0.2 µm) in this study. Further investigation revealed that the effects of pore size on retention could be correlated to the particle size distribution in the cell culture broth. A hollow fiber with a larger pore size (>0.5 µm) may reduce protein retention when small particles (approximately 0.01-0.2 µm in diameter) are dominant in the culture. However, if majority of the particles are larger than 0.2 µm in diameter, hollow fiber with smaller pore sizes (0.2 µm) could be a solution to reducing product retention. Alternatively, process optimization may modulate particle size distribution towards reduced production retention with selected ATF hollow fibers. This study for the first time highlights the importance of matching proper pore sizes of hollow fibers with the cell culture particles distribution and offers methods to reducing product retention and ATF column clogging in perfusion cell cultures. KEY POINTS: The material of ATF column could impact product retention during perfusion culture. Higher harvest flowrate and ATF exchange rate increased product retention. Matching culture particle size and ATF pore size is critical for retention modulation.