What contributes to successful transfection?

Cell line health and viability, nucleic acid quality, transfection reagents, duration of transfection, and serum requirements all affect the success of transfection. Cells must be healthy, uncontaminated, and in the correct growth phase and density (typically 40-80% confluent for chemical methods). Nucleic acid quality (high purity, low endotoxin levels) and appropriate reagent concentration are also crucial for optimal outcomes.

Is it possible to predict the transfectability of a specific type of cell?

No, predicting transfection efficiency based solely on cell type is not possible. The interactions between cell properties and transfection reagents are complex. However, it is empirically known that some cell types, such as primary cells, quiescent cells, and suspension cells, are generally difficult to transfect.

What impact does the type and quality of DNA or RNA have on transfection success?

High-purity genetic material consistently leads to better transfection outcomes. Endotoxin contamination, introduced during bacterial processing, must be removed as it can trigger the innate immune system and hinder transfection. Additionally, promoter characteristics, gene properties affecting cell physiology or survival, and the nucleic acid's size and structure all influence transfection success.

How is Transfection Different from Transformation and Transduction?

Transfection refers to the introduction of foreign nucleic acids into cells, typically by non-viral methods, especially in animal cells. Transformation describes the same process in plant, fungal, and bacterial cells, but is avoided in animal cell biology due to its association with 'malignant transformation' in cancer. Transduction refers to the viral-mediated transfer of nucleic acids into cells.

Supporting a Maturing Modality: Large-Volume Manufacturing of Mesenchymal Stem Cells

Large-Volume Manufacturing of Mesenchymal Stem Cells

Brian Gazaille with Khang Luu, and Irana Coletti Malaspina

This article was originally published in BioProcess International's Manufacturing Cell & Gene Therapies featured report, April 2026.

Mesenchymal stem/stromal cells (MSCs) are making headway in biopharmaceutical development pipelines. Khang Luu and Irana Coletti Malaspina (both field application scientists at Corning Life Sciences) explain that such cells were used initially to tamp down immune responses in cases such as graft-versus-host disease (GvHD) and osteoarthritis. Researchers have since discovered that MSCs differentiate into several therapeutically valuable cell types, secrete molecules that contribute to tissue repair, and exhibit anti-inflammatory properties that enable safe allogeneic administration. Thus, developers are investigating MSCs and MSC-derived exosomes for treatment of chronic wounds and inflammatory diseases. MSCs also naturally infiltrate tumors, opening up possibilities for engineering MSCs to deliver therapies to cancer cells.

Despite clinical progress, large-volume MSC manufacturing remains difficult. I corresponded with Luu and Malaspina to learn about efforts to increase the scalability of MSC production. Their combined responses are below. With multiple MSC candidates knocking on the door to commercialization, the industry needs advanced cell-culture technologies that will support the modality’s maturation.

Improving MSC Yield and Quality

My impression is that producing large quantities of MSCs remains challenging. What factors hamper MSC production?

Historically, biological constraints have limited large-scale production.

Suboptimal culture components — such as low-grade media, reagents, and consumables — can impair MSC growth from the outset. Cells expanded with nonoptimal materials often show reduced proliferative capacity and inferior overall performance in subsequent passages. Poor environmental conditions also can cause MSCs to deviate from their intended lineage, leading to unwanted differentiation and compromising product quality and consistency.

Production techniques must be tailored to a facility’s infrastructure, staffing levels, scale of operation, and end goals, whether the objective is producing MSCs or harvesting their secreted factors. Maximizing yield depends heavily on workflow optimization and equipment that can process cells gently enough to preserve their physical integrity, biological characteristics, and potency.

Long-term stability raises difficulties, too. MSCs are highly sensitive to storage conditions, temperature fluctuations, and handling, all of which can compromise their viability and therapeutic function.

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What quality markers should be monitored during production, and why is it difficult to preserve those throughout manufacturing?

Guidelines from the International Society for Cell & Gene Therapy (ISCT) clearly define required MSC surface markers and biological functions. Such characteristics can be fully assessed only at the end of production, so release testing is required to confirm MSC identity, purity, and functional quality. Moreover, each batch must undergo quality control (QC) testing after production to ensure that it meets specifications.

During process validation, critical quality attributes (CQAs) must be established in alignment with regulatory expectations. Manufacturers must demonstrate that a process will produce MSCs that consistently meet release criteria despite the inherent biological variability of cell-based products.

A major impediment is the current lack of validated secreted biomarkers that can be analyzed from a culture environment to predict final MSC quality.

Throughout manufacturing, MSCs remain sensitive to changes in their culture environments, so they can differentiate inadvertently into non–stem-cell phenotypes if conditions are not carefully maintained.

Many developers apply a quality by design (QbD) approach to address such challenges. A QbD strategy emphasizes designing and optimizing a robust production process — e.g., using a reliable cell source and high-quality reagents and culture materials — to increase the likelihood that MSCs will meet quality requirements during final QC and release testing.

What parameters should sponsors prioritize during MSC expansion?

Manufacturers should emphasize conditions that support consistent cell growth, preserve stem-like characteristics, and minimize variability across scale-up steps. Expansion typically follows a seed-train approach, moving from small vessels such as T flasks to larger systems such as Corning HYPERStack vessels and CellCube systems. At each transition, scientists should monitor contamination, cell morphology, and yield; a well-controlled process should produce relatively stable yields across scales, with only minor variations.

Significant fluctuations often signal underlying issues with culture health or process stability.

A well-controlled culture environment is equally important. Manufacturers should use reliable, high-quality consumables and reagents, ensuring that supplements are fresh and consistent in grade. When possible, media formulations should be designed specifically for MSC expansion. Sponsors also should limit passage numbers to preserve MSC “stemness.” Doing so requires a well-qualified master cell bank (MCB) and monitoring of CQAs throughout expansion.

To maintain cell viability and potency throughout manufacturing, downstream-processing and fill–finish equipment must handle cells gently. Moreover, MSC-aggregate formation should be managed carefully through appropriate process controls. Factors such as confluency at harvest, environmental conditions, media composition, harvest reagents, and agitation parameters all play a role in preventing aggregation, which can diminish cell quality and performance.

What equipment formats can help to address MSC scalability concerns?

Platforms such as Corning CellSTACK culture chambers, HYPERStack vessels, and CellCube systems can be used for both scale-out and scale-up strategies depending on how your process begins and what your production goals are.

Such systems provide reliable, well-characterized environments for MSC culture and have been adopted widely throughout the industry because they meet diverse application and product needs.

MSC yield requirements can differ substantially depending on route of administration, dosing frequency, disease indication, and patient population. Such variables determine required cell numbers per batch and thus whether to scale up or out. Companies also must ensure that CQAs remain stable throughout production. Scale-up approaches (e.g., expanding into CellCube systems) are often a good choice for products that require large quantities of cells and/or long-term storage. When small batches are needed at high frequencies for immediate use, scale-out approaches (with Corning HYPERStack vessels or CellSTACK culture chambers) might be the better option.

Amid variability in demand, modular technologies (including CellSTACK chambers, HYPERStack vessels, and CellCube systems) can provide flexible scaling options by enabling adjustment of total culture surface area while maintaining a consistent growth environment. Such flexibility supports predictable, reproducible MSC expansion and helps manufacturers align their production strategies with therapeutic, regulatory, and operational requirements.

How much does vessel-surface treatment influence MSC expansion?

It has a significant impact because MSCs are adherent cells. Depending on your culture conditions, particularly when using serum-free media, specialized technologies such as Corning’s CellBIND surface might be necessary to ensuring robust adhesion and optimal growth. When MSCs must be grown in three-dimensional spheroid formats, such as for enhanced exosome production, ultralow-attachment (ULA) surfaces are required to prevent cell adherence. Even among standard treatments, variations in surface chemistry and manufacturing among brands can lead to differences in attachment efficiency, affecting MSC growth rates and expansion performance.

Are you aware of promising developments in the MSC field?

Japanese regulatory authorities recently approved two therapies derived from induced pluripotent stem cells (iPSCs). In addition to marking global firsts for the field, those approvals signal growing regulatory confidence in advanced cell based therapeutics and highlight increasing momentum behind next-generation stem-cell technologies. Such progress not only underscores the maturation of the cell-therapy industry, but also provides important insights for future development and commercialization pathways related to MSC-based therapies and other regenerative treatments.

Brian Gazaille is managing editor of BPI; brian.gazaille@informa.com. Khang Luu, PhD, is a field applications scientist for Southeast Asia, and Irana Coletti Malaspina, MSC, is a field application scientist for South America from Corning Life Sciences. Corning, CellCube, CellBIND, CellSTACK, HYPERFlask, and HYPERStack are all trademarks of Corning Incorporated.