June 10, 2024
2:26 pm

Applications of Single-Cell Seeding in Biomedical Research

By Katrin Raute

Product Manager

Applications of Single-Cell Seeding in Biomedical Research

Single-cell seeding is revolutionizing biomedical research, offering unprecedented disease insights and treatment options in various fields. Its applications span a range of disease states and drug development phases. Regenerative medicine using induced pluripotent stem cells (iPSCs) benefits immensely, as single genetically modified stem cells can be scaled to regenerate disease tissues. In oncology, single-cell seeding facilitates a better understanding of tumor heterogeneity and the development of targeted treatments. Single-cell seeding is a cornerstone technology driving innovation and improving patient outcomes in modern medicine. Here, we will discuss single-cell seeding applications in translational biomedical research and highlight how new technologies, like the UP.SIGHT from CYTENA, help to overcome complex challenges in this field.

Regenerative Medicine

Regeneration of diseased or dead tissue has been a promise of stem cell research since its inception. Single-cell seeding applications in regenerative medicine allow single stem cells to be isolated and scaled up to treat many diseases and conditions.

iPSCs

The isolation and manipulation of iPSCs is a cornerstone of regenerative medicine¹. These cells can be taken from an individual and differentiated into a cell type that can help replace or grow missing or damaged tissue. Advances in genetic engineering, particularly with the Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) technology, allow donor iPSCs to be altered to overcome issues with rejection. Furthermore, researchers can culture specific stem cells using single-cell dispensing to gain insights into their function and behavior under different conditions.

Accurate single-cell seeding is crucial here for establishing a traceable clonal population that will produce consistent and replicable differentiation states under a given treatment (Fig. 1). Thus, single-cell seeding applications in regenerative medicine impact a wide variety of processes, including:

Cardiac Regeneration
Wound Healing
Spinal Cord Injuries
Liver Regeneration
Retinal Regeneration
Bone and Cartilage Repair

Figure 1. Single stem cells can be programmed and scaled to regenerate many bodily tissues making them a uniquely powerful therapeutic strategy.

In-vitro Modeling

Many research areas lack models replicating the complexity of multi-tissue structures like organs. Indeed, many research areas use a single cell line that does not accurately replicate tissue, let alone the complete organ. Single seeding of stem cells allows organoids (mini-organs) to be grown in culture, and the number of specific cell types to be adjusted, providing a far better model for understanding cellular function or testing the response to different therapies².

Oncology: Understanding Tumor Heterogeneity

Tumors are typically complex mixtures of different cell types with various capabilities and vulnerabilities³. In oncology, single-cell seeding applications allow for better characterization of the myriad of cells within a tumor mass and provide a platform for developing biological treatments like antibodies and cell-based therapies (Fig. 2).

Figure 2. Single-cell seeding allows researchers to hone in on specific cancer cells and gain strategic insights into their vulnerabilities.

Cancer Stem Cells

A central challenge in oncology is treatment resistance. While many cancer cells succumb to treatment, some cells survive, proliferate, and form tumors years later. These are known as cancer stem cells (CSCs)⁴. Recurrent cancers are typically more aggressive than the original tumor⁵, so the characterization of CSCs via single-cell seeding allows scientists to understand how they resist treatment and develop strategies to resensitize them.

Immune Component

Many tumors possess an immune cell component that helps the cancer resist therapy. Several treatments aim to rearm the immune system to attack the tumor instead of aiding it. Examples include antibody-based therapies like programmed cell-death 1 inhibitors, which prevent cancer cells from “switching off” immune cells⁶. These therapies require single-cell seeding to provide verifiable, high-yield cell lines that produce antibodies⁷.

Metastasis

Metastasis is a leading cause of cancer-associated mortality. However, understanding which cells are capable of metastasis (and why) remains a largely open question in cancer research. Circulating tumor cells (CTCs) found in the bloodstream offer a unique window for studying and characterizing cells as they undergo metastasis⁸. Single-cell seeding allows these cells to be cultured, thus eliminating the potential noise of other cells, either from the primary tumor or blood cells⁹.

Therapy Development

While single-cell seeding allows us to gain greater insights into different diseases, it also provides novel ways to develop treatments against them. We’ve already examined how single-cell seeding helps in the early phases of translational research; now, let’s examine how this technology helps transform disease insights into real therapies for patients.

Drug Screening

Single-cell seeding allows drugs to be screened against specific cell populations to assess the drug’s ability to kill a target cell or to determine its toxicity on non-target cells. This allows researchers to fine-tune dosage, limit toxicity, and enhance efficacy against the target population.

Therapy Production

Cells can be used as factories to produce biologic-based therapies or as therapies in their own right. Both cases require accurate single-cell seeding at the beginning of the therapy production process to ensure purity and functionality. Regulatory bodies require robust traceability of cell lines used as or to produce therapies¹⁰. Without accurate single-cell seeding, this becomes increasingly difficult, leading to delayed drug discovery timelines and significant loss of resources.

The Challenges of Single-Cell Seeding in Biomedical Research

Single-cell seeding is a complicated process that must be performed accurately and consistently. Applying this technique becomes even more challenging when we consider the complexity of the diseases and the research question it helps to address.

Biological Challenges

Unlike pharmaceutical drugs, cells are living entities and are, therefore, impacted by stressors that hinder their growth and reduce their functionality. Maintaining sufficient growing conditions while ensuring that cells are not exposed to stresses associated with handling is a significant challenge. For example, iPSCs are sensitive to shear stresses common to isolation techniques like FACS.

Technical Challenges

Single-cell seeding requires accurate dispensing of small amounts of liquid, ensuring a single cell is added to a single well. Manual techniques to achieve this are laborious and prone to errors, meaning higher chances of contamination and lower chances of achieving clonality. Manual monitoring of clones is also time-consuming and introduces challenges for data management and traceability, forcing researchers to spend hours at a microscope instead of analyzing data and brainstorming new ideas.

These challenges significantly impact research and drug development timescales, resulting in business failure and a massive waste of time and resources.

The UP.SIGHT All-in-One Single-Cell Dispenser and Imager

This novel technology from CYTENA uses automation and microfluidics¹¹ to ensure fragile cells like iPSCs are exposed to less shear stress and achieve higher recovery rates. The UP.SIGHT’s advanced imaging technology ensures clonality and makes it easy for scientists to select and scale up high performers (Fig. 3). In short, the UP.SIGHT provides a one-stop solution to various biological and technical challenges in single-cell seeding.

Figure 3. The UP.SIGHT’s imaging capabilities allow scientists to track the growth of adherent and non-adherent cells with the help of C.STUDIO software.

Conclusion and Future Directions

Single-cell seeding is a transformative technology in biomedical research, driving significant advancements in disease understanding and therapy development in regenerative medicine and beyond. New technologies that integrate non-coding scheduling software and automation for image and data analysis will help streamline workflows, while novel biomaterials will likely improve cell functionality and recovery. Advances in our technological capabilities and understanding of disease ensure that single-cell seeding will continue revolutionizing biomedical research, paving the way for innovative treatments and more personalized medicine approaches.

Cutting-edge equipment like the UP.SIGHT from CYTENA allows scientists to establish themselves as pioneers in this rapidly evolving area. Contact our team to learn more about the UP.SIGHT or book a demo to begin your journey with CYTENA.

References

Nicholson MW, Ting CY, Chan DZH, et al. Utility of iPSC-Derived Cells for Disease Modeling, Drug Development, and Cell Therapy. Cells. 2022;11(11):1853. doi:10.3390/cells11111853
Kim J, Koo BK, Knoblich JA. Human organoids: model systems for human biology and medicine. Nat Rev Mol Cell Biol. 2020;21(10):571-584. doi:10.1038/s41580-020-0259-3
Hausser J, Alon U. Tumour heterogeneity and the evolutionary trade-offs of cancer. Nat Rev Cancer. 2020;20(4):247-257. doi:10.1038/s41568-020-0241-6
Batlle E, Clevers H. Cancer stem cells revisited. Nat Med. 2017;23(10):1124-1134. doi:10.1038/nm.4409
Damen MPF, Van Rheenen J, Scheele CLGJ. Targeting dormant tumor cells to prevent cancer recurrence. The FEBS Journal. 2021;288(21):6286-6303. doi:10.1111/febs.15626
Sharma P, Goswami S, Raychaudhuri D, et al. Immune checkpoint therapy-current perspectives and future directions. Cell. 2023;186(8):1652-1669. doi:10.1016/j.cell.2023.03.006
Parray HA, Shukla S, Samal S, et al. Hybridoma technology a versatile method for isolation of monoclonal antibodies, its applicability across species, limitations, advancement and future perspectives. Int Immunopharmacol. 2020;85:106639. doi:10.1016/j.intimp.2020.106639
Ring A, Nguyen-Sträuli BD, Wicki A, Aceto N. Biology, vulnerabilities and clinical applications of circulating tumour cells. Nat Rev Cancer. 2023;23(2):95-111. doi:10.1038/s41568-022-00536-4
Teng T, Yu M. Establishing Single-Cell Clones from In Vitro-Cultured Circulating Tumor Cells. In: Gužvić M, ed. Single Cell Analysis. Vol 2752. Methods in Molecular Biology. Springer US; 2024:119-126. doi:10.1007/978-1-0716-3621-3_8
Welch JT, Arden NS. Considering “clonality”: A regulatory perspective on the importance of the clonal derivation of mammalian cell banks in biopharmaceutical development. Biologicals. 2019;62:16-21. doi:10.1016/j.biologicals.2019.09.006
Liu D, Sun M, Zhang J, et al. Single-cell droplet microfluidics for biomedical applications. Analyst. 2022;147(11):2294-2316. doi:10.1039/D1AN02321G