James L Sherley, MD, PhD, explores how a new method for quantifying stem cells in mesenchymal tissue samples can lead to better drug development for stem cell therapies.  

Mesenchymal tissue cell samples are considered incredibly promising for the development of new stem cell therapies. These samples can be based on bone marrow, umbilical cord tissue, adipose tissue, and other common sources. But there’s a problem with mesenchymal tissue cell preparations: they’re highly heterogeneous, containing the target stem cells mixed with many other cells that aren’t useful for therapy production. And as these samples are cultured, the fraction of stem cells can decrease over time, making it that much harder to use these samples as the basis for new therapies. 

These issues contribute to uncertainty in the development and manufacture of stem cell therapies, incorrect dosing for patients, and wasted stem cell resources, among other problems. With so much potential for treating a host of diseases that have previously been incurable, stem cell therapies and the regenerative medicine field working to bring them to patients deserve a more rigorous and robust approach. 

Recently, scientists made new discoveries about the stem cell expansion process that have direct implications for stem cell therapy development¹. Those efforts have advanced therapeutic discovery, development, and manufacturing in two key ways. First, a novel method known as kinetic stem cell counting has for the first time made it possible to quantify the fraction of stem cells present in a mesenchymal tissue sample — all with greater accuracy and speed than any conventional counting technique. This method also gave rise to the second advance: a new understanding of the biological mechanism that explains why some stem cell fractions increase or stay the same in culture while others decline over time. 

While these innovations have immediate utility for stem cell therapy development, they will also support improvements to biomanufacturing, lab-cultured food production, and even reproducibility in basic research. 

Kinetic stem cell counting 

Prior to this new research, there was only one technique for differentially counting any type of tissue stem cell — and those were haematopoietic stem cells, not mesenchymal stem cells. That lone technique is the dreaded SCID mouse repopulating cell assay. It involves injecting up to 40 mice in a very expensive process, caring for them for more than four months, and then analysing their tissues and organs. After this cumbersome, costly, and time-intensive operation, the resulting count is neither reliable nor terribly accurate. Alternative counting methods, such as using the CD34 marker or colony-forming unit assays, are unable to distinguish between the stem cells needed for therapeutic use and progenitor cells that lack curative properties. The counts they generate are therefore falsely high. 

Kinetic stem cell counting, on the other hand, is rapid and accurate. Stem cells create more stem cells through symmetric division, but produce committed progenitor cells or differentiated cells through asymmetric division. By using models of the different cell kinetics involved in each of these processes, the new counting method can quantify the stem cell fraction of a cultured tissue cell sample. The technique involves a computational simulation, reducing the risk of human operator error in the quantification process. 

Being able to perform differential counts of tissue stem cells opens entirely new opportunities in the discovery and development of stem cell therapies. Kinetic stem cell counting makes it possible to determine the different fractions of stem cells as well as their progenitor cells and arrested cells in a tissue cell preparation; it can also reveal how those fractions change over time in cell culture. The approach could be used, for example, to test drugs for toxicity to stem cells in culture. This could help flag candidates that would be more likely to fail late in clinical trials, providing crucial information much earlier in the development process so resources could be allocated to stronger candidates. 

Overall, the capabilities of kinetic stem cell counting are particularly useful for optimising the yield of tissue stem cells used in drug discovery research, drug candidate evaluations, and transplant therapies. It can also support much-needed process improvement for stem cell production techniques that are a cornerstone of discovery, development, and manufacturing methods related to the use of tissue stem cells. 

Tackling the turnover unit 

With a reliable and rapid means of counting stem cells, it is now possible to figure out the stem cell fraction and monitor how it changes over time. In the past, scientists have noted that the stem cell fraction can increase, decrease, or remain the same over time in cell culture². But how it changed, and how it could be prevented from decreasing when those cells need to be used for medical purposes, has been a mystery. 

Thanks to accurate counting, those mysteries have now been solved. The answer came from the stem cell turnover unit, or the number of cells representing the committed cell lineage that has been produced by the tissue stem cell. As it turns out, mesenchymal tissue cell samples with more stable stem cell fractions over time have smaller turnover units, while samples with decreasing stem cell fractions have substantially larger turnover units. The concept behind this is straightforward: smaller turnover units reduce the production of committed progenitor cells, allowing for greater expansion of tissue stem cells. 

This finding would be largely academic if it weren’t for one critical factor: the availability of agents that can change the turnover unit size of stem cells³ Cellular purines such as xanthosine have been shown to suppress asymmetric cell kinetics, increasing the yield of stem cells in a tissue cell sample. The latest discoveries make clear that xanthosine has this effect because it stops the stem cells from making turnover units and boosts the number of stem cells they produce instead. 

With this new insight, biopharma scientists can now improve the stem cell yield of their mesenchymal tissue cell samples. This has important ramifications for manufacturing stem cell therapies. Historically, expanding these samples in culture has led to the well-known loss of stem cell function, limiting the success of stem cell therapies. Some of that loss of function has been linked to a decline in stem cell fraction — a problem that can now be addressed with the use of appropriate agents and kinetic stem cell counting. 

Having more control over stem cell production in these tissue cell samples will lead to significant advantages for biopharma teams. Changes to the stem cell fraction in samples expanded for therapeutic use are expected to become a critical quality attribute for stem cell treatments. In addition, manipulating stem cell fraction could help to ensure that patients get the appropriate therapeutic dose. Today, too many patients get a lower-than-intended dose, which depresses therapeutic response, or a higher-than-necessary dose, which wastes valuable stem cell resources that could have been used for another patient. 

Stem cell counting in action 

While kinetic stem cell counting is still quite new, the scientific basis for it has already contributed to new tools designed to make accurate cell counts more accessible for scientists throughout drug discovery and development. For example, the global standards organisation ASTM International released a new standard last year that provides detailed guidance for how to calculate cell proliferation rate⁴. This F3716 standard was designed for use with serially maintained cultures and can be used for regenerative medicine programmes such as stem cell therapy development. Work is currently underway to design a follow-up standard specific to differential stem cell counting⁵. 

Late last year, the FDA recognised the F3716 standard, incorporating it into its industry guidance list of voluntary consensus standards related to regenerative medicine therapies⁶. The standards recognition programme is overseen by the Center for Biologics Evaluation and Research in order to support the development of new regenerative medicines. 

In addition, scientists have developed and made freely available online calculators aimed at helping researchers measure cell proliferation rates more accurately⁷. This information is crucial for the accurate quantification of tissue stem cells in culture. The calculators use simple inputs, including cell number and time between passages, to tally cell proliferation rates and other key metrics. 

What’s next? 

Stem cell therapies are an exciting new class of treatments, with tremendous potential to reverse damage, mitigate serious symptoms, and even cure previously intractable diseases. But several technical limitations — including the inability to produce accurate counts of these cells in a cultured sample and declining function of stem cells expanded in culture — have hindered progress in regenerative medicine. The development of kinetic stem cell counting and its contributions to a new understanding of the influence of turnover unit size on stem cell production are key advancements that could ultimately help bring stem cell therapies to more patients in need. 

References   

1. Sherley JL, Cao C, Sommer C,et al. The Size of Human Mesenchymal Tissue Stem Cell Turnover Units is a Major Determinant for Maintaining High Stem Cell Fractions With Serial Culture Expansion. Stem Cells Int. 2026 ;2026:1216876. 
2. Chopra H, Cao C, Sommer C,et al. Quantification of the Culture Stability of Stem Cell Fractions from Oral-Derived, Human Mesenchymal Stem Cell Preparations: A Significant Step toward the Clinical Translation of Cell Therapies. Cells. 2023;12(23):2703. 
3. ParéJ-F, Sherley JL. Ex vivo Expansion of Human Pancreatic Distributed Stem Cells by Suppression of Asymmetric Cell Kinetics (SACK). J Stem Cell Res & Therapy 2013;3:149. 

4. ASTM International.Standard Test Method for Cumulative Population Doubling Analysis of the Proliferation of Vertebrate Tissue Cell Preparations.https://store.astm.org/f3716-25.html. Last updated Feb 21, 2025. Accessed Feb 13, 2026. 

6. ASTM International.New Test Method for Differential quantification of stem cells in samples of isolated postnatal vertebrate tissue cells.https://www.astm.org/membership-participation/technical-committees/workitems/workitem-wk95346. June 2025. Accessed Feb 13, 2026.
7. USFood and Drug Administration.Standards Development for Regenerative Medicine Therapies. https://www.fda.gov/vaccines-blood-biologics/standards-development-regenerative-medicine-therapies. Last updated Jan 27, 2026. Accessed Feb 13, 2026. 
8. Asymmetrex. Stem Cell Counting Center.https://asymmetrex.co/stem-cell-counting-center/. Accessed Feb 13, 2026. 

About the author 

James L Sherley, MD, PhD, spent decades as a cancer cell molecular biologist and stem cell scientist in organisations including MIT, Fox Chase Cancer Center, and Boston Biomedical Research Institute. His company, Asymmetrex, is developing new methods and devices for differential stem cell counting to support better science and medicine.